Bovine Trichomoniasis. A. Yule*, S.Z. Skirrowtand R.H. BonDurantf. Although virtually unknown in Europe since the widespread adoption of artificial insemi-.
373
Porositology Today, vol. 5, no. I2, I989
Bovine Trichomoniasis A. Yule*, S.Z. Skirrowtand
R.H. BonDurantf
Although virtually unknown in Europe since the widespread adoption of artificial insemination (AI), infection by (thesexually transmittedprotozoan parasite Tritrichomonas foetus (Fig. I) resultsin substantial economic lossesthroughout the major cattle-rearingareas of the world where natural breeding is relied upon. Infection by T. foetus is increasingly recognized as a significant cause of ,bovine infertility. In this review, Alex Yule, Susan Skirrow and Robert BonDurant summarize the current knowledge of bovine trichomoniasis and the problems of diagnosis and controlof thiseconomicallyimportantdisease.
The link between bovine trichomoniasis and infertility was well established by the middle of this century (early research is summarized in Morgan’s 1946 monograph)‘, but the importance of the disease was somewhat overshadowed by brucellosis. The effective control ofBrucella abortusinfection and that of another venereally transmitted pathogen, Campylobacterfetus subspecies venerealis, has shown that T. foetus infection is also a significant cause of bovine reproductive failure. Australian surveys, based on the culture of post-mortem samples and samples from live animals found a prevalence in , and a recent beef herds of 40-650Y02y3 survey of randomly sellected herds suggested a prevalence of about 14% in Californian beef cattle (RX BonDurant et al., unpublished). Reports of within-herd prevalence of trichomoniasis in bulls r,ange from 26.4% in South Africa4 to 30.6-50% in Australia5a6and 5.8-38.5% in CNifornia7~8. Few recent figures are available for prevalence in cows. Australian surveys have shown levels of infection of 8.40/b2,and similarly, 10.7% of cows were found to be infected in a large Californian dairy herd’. However, in a simulated natural infection, Clark et al. lo established that 42.5% of cows exposed to T. foetus-infected bulls were infected after the first year.
predicted a reduction of 5% in the return per beef cow, when the prevalence of T. foetus reached 20% among bulls, but if this prevalence doubled, the economic return from each cow would drop by 35%“. Pathogenesis and pathology Transmission of T. foetus is almost
exclusively by coitus. The parasite can survive in whole or diluted semen not only at 5°C but also after semen cryopreservation12y13.Infection through AI is rare, although mechanical transmission via insemination instruments or vaginal palpation has been known to occur”. Infection is confined to the reproductive tract, and in bulls the preputial cavity and, less commonly, the urethral orifice14 are preferred by this parasite. Organisms are only found on mucosal surfaces, in secretions and gland lumina14~15, and in males there is generally no gross indication of infection and only minor infiltrations of neutrophils, macrophages and lymphocytes are seen on microscopic examination14. The prevalence of infection in bulls
Economic significance
The economic consequences of T. foetus are poorly known and few analyses have been made. In 1988, McCool estimated that 25 500 calves were lost each year in the Victoria River district of northern Australia3, indicating that the annual financial burden imposed by trichomoniasis could be in the order of tens of millions of US dollars. Losses due to reduced milk and calf production in a Californian dairy herd were calculated as US$665 per infected cow9. A computer simulation @,989, Elsewr Scmce Publ,shers Ltd. (UK) 0 l65’%4707189l$O2 CO
Fig.
I. Tritrichomonas
foetus.
*Department of Medical Parasitology London School of Hygiene and Tropical Medicine Keppel Street London WC I E 7HT. UK tMoe Veterinary Clinic 3 I Lloyd Street, Moe Victoria 3825 Australia *Department of Veterinary Reproduction School of Veterlnaty Medicine University of California Davis CA 956 16. USA
Parasitology
374
seems to increase with age3>8,possibly due to the deepening of crypts in the preputial epithelium, which provide an ideal niche for the parasite, but the number of matings during the lifetime of each animal must obviously also play a part. Field evidence suggests that infected bulls, of 4 years and older, tend to remain carriers for life’, although younger bulls are not exempt from infection7’8. Differences in the susceptibility of different breeds have been suggested 16y17,but have not been unequivocably determined. Similar infection rates have been seen in Bos tam-usand Bos indicus breeds’. While the pathogenesis of trichomoniasis in the cow is still poorly known, the progress of infection appears to be rapid, and all regions of the reproductive tract are colonized within two weeks of infection”. The numbers of parasites present in the cervico-vaginal mucus fluctuate during the oestrus cycle, and the largest numbers are seen a few days before oestrus”. Initially, the uterus was thought to be the main site of infection17’20, but several studies of naturally infected cows indicate that the OScervix is the preferred site18. Infection is self-limiting in females, with clearance of parasites from the reproductive tract occurring after approximately 95 days14; however, the phenomenon of the carrier cow has been recently confirmed*‘. While carriers are rare, the ability to maintain infection throughout pregnancy, and for six to nine weeks postpartum, provides a source of reinfection for bulls. T. foetus only occasionally causes mild vaginitis with mucopurulent discharge’* and does not prevent conception14. Most pregnancies are lost about 17 days after conception when maternal recognition has occurred, but embryonic death can occur at any time up to the fifth month of gestation (cattle have a gestation period of nine months)**. Little pathology is seen until about 50 days after infection. Acute to chronic mild inIlammation develops, characterized by accumulations of polymorphonuclear granulocytes, macrophages, lymphocytes and small numbers of plasma cells throughout the reproductive tract15, even though the parasite is confined to the epithelial cell surface of the endometrium and placenta. Mild placental lesions associated with invasion of the chorionic epithelium have been described in cows aborting after the first three months of pregnancy23. Following foetal
Today, vol. 5, no. 12. I989
loss, there is a gradual process of repair leading to restoration of a fertile uterus two to six months after infection16, although retention of the corpus luteurn or a macerated foetus can give rise to pyometra and chronic inflammation, which occasionally result in permanent infertility12>24. There is a paucity of research concerning the mechanism(s) by which T. foetus exerts its effect on the conceptus. Experimental evidence suggests a role for tumour necrosis factor (TNF) in malariainduced abortion25, and lymphokinemediated cytotoxicity may be important in bovine trichomoniasis. Alternatively, the parasite may directly affect the embryonic or foetal tissues by the production of as yet uncharacterized cytotoxins. T. foetus secretes several low molecular weight cysteine proteinases, of which two, with apparent molecular masses of 21.6 and 19.8 kDa have been found in the cervico-vaginal mucus of artificially infected beef heifers (A. Yule et al., unpublished). Immunity to infection
Very little recent work has been done on immunity to T. foetus but the early work has been comprehensively reviewed by Honigbergz6. Recent experimental work has not detected significant serum levels of T. foetus-specific IgA, IgG and IgM using an enzyme-linked immunosorbent assay (ELISA)27. Hypersensitivity reactions have been shown in infected cows on intradermal challenge” but the role of IgE in the pathogenesis of trichomoniasis remains unknown. Although circulating antibodies appear to offer little protection against reinfection*‘, an infected cow is capable of clearing parasites from the reproductive tract and does become temporarily refractive to reinfection’*. Simultaneous clearance of T. foetus from the uterus, cervix and vagina was observed in a recent experimental study18 and may be attributable to local antibodies to T. foetus as suggested by the earlier work26. Vaginal and cervical IgA and IgGl responses have been found to increase about eight weeks post-infection, whereas increases in uterine IgA and IgGl occurred at approximately week 1127. Very little IgM was detected, and IgG2 appeared only transiently, increasing at about week 14 in the vagina and week 11 in the cervix. The specificities of the IgA and IgGl responses were inconsistent throughout infection, and a variety of antigens ranging from 20 to 200 kDa were
ParasitologyToday,vol. 5, no. 12, I989
detected, although parasite clearance appeared to be correlated to the recognition of high molecular weight antigens (S.Z. Skirrow and FLH. BonDurant, unpublished). There is scant recent evidence for cellmediated immunity in bovine trichomoniasis but lymphocytes and polymorphs from mice immunized with live T. foetus have been shown to kill parasites on contact in vitro2*. Phagocytosis of T. foetus by pulmonary macroph.ages has been observed in aborted bovine foetuses23.
infected was estimated as only 59%“. Immediate inoculation of culture medium is not always possible in the field and transport media are valuable in maintaining parasite viability7. The application of immunological methods to the detection of T. foetus infection has so far met with only limited success and again little work has been done since the 1940s. The direct detection of antigen in clinical samples has proved far more difficult in the case of T. foetus than with its counterpart in humans, Trichomonas vaginalis. The development of a Diagnosis of T. foetus infection sensitive test has been complicated by The presence of trichomoniasis within a cross-reactivity problems and by the low beef herd is initially indicated by lower z;$ of antigen present during infecthan expected pregnancy rates and an extended calving seaso:n. Early abortion and occasional pyometra may occur, Control of infection Various topical treatments have been although the prevalence of these symptoms is often less than 5% in newly advocated for the elimination of infection infected herds22. As trichomoniasis is so in bulls (eg. Ref. 33) but are not consissimilar to Campylobacter fetus venerealis tently effective and involve repeated infection, the absence of this latter patho- applications to often increasingly relucgen needs to be confirmed as does the tant patients. The therapeutic value of several 5absence of other causes of reproductive failure, such as lowered bull fertility and nitroimidazole compounds approved in the USA for the treatment of Histomonas inadequate nutrition. Since infection is inapparent in the bull, mekagridis (blackhead disease) in turkeys and mild vaginitis is only occasionally has also been established for T. foetus. found in some COWS~~,~~:, a definitive diag- Multiple oral doses or intravenous adminnosis can only be made by direct obser- istration of dimetridazole are effective but vation of the parasite in preputial or can result in serious side-effects, from vaginal secretions and in placental and anorexia to respiratory difficulty, ataxia abomasal fluids from aborted foetuses, or and collapse7y34. The intramuscular administration of ipronidazole is free of sideby culturing it in one of sleveral media29. If possible, bulls are sexually rested for effects and haematological and serum at least one week before sample collection chemistry values remain within normal to maximize the number of organisms in ranges in treated animals (R.H. Bonthe preputial cavity 13. A variety of tech- Durant, S.Z. Skirrow and B. Norman, niques have been described for sample unpublished)3s, although sterile abscesses collection7J6’22’30. T. jbetus are recog- may form at the injection site22. Multiple nized microscopically in clinical samples doses are more effective than single doses’ may be further by their jerky, rolling progression but the and effectiveness number of trichomonacls found in pre- enhanced by prior or simultaneous putial secretions is highly variable (200 to administration of antibiotics to eliminate > 80 000/m1)‘2, and direct examination is the bacterial degradation of this drug in approximately 25% less sensitive than the preputial cavity3(j. Inadequate dosage culture31 (S.Z. Skirrow and R.H. Bon- may render infection undetectable for up to four weeks after treatment’ and thus Durant, unpublished). Culture of a single sample has been culture after this time is essential to concalculated to give only an 80-90% chance firm the complete elimination of T. foetus. of obtaining a positive result from a bull Resistance to S-nitroimidazole therapy known to be infected7>*, and the culture of does occur17 and, as with metronidazoleat least three successive samples is resistant T. vaginaZis37, also appears to required to minimize the risk of obtaining confer resistance to other 5nitroimidazoles38. a falsely negative result. Unfortunately, Although undoubtedly effective, 5culture sensitivity is even lower in cows; the chance of obtaining a positive culture nitroimidazole treatment is costly. Moreof a single sample from a cow known to be over, the use of these compounds is not
375
376
Parasrtology Today, vol. 5,
approved by the United States Food and Drug Administration and ipronidazole has recently been declared a suspect carcinogen and withdrawn from the market. Therefore, in the western USA at least, herd management is the sole means of preventing and controlling infection. Current strategies for the control of T. foetus in beef and dairy cattle are reviewed in detail by Ball et al. l7 and are based on the prevention of transmission, the elimination of infected animals and the prevention of reintroduction of infection into a clean herd. Control may be complicated by other management practices such as the sharing of grazing land or the leasing of bullsZ2. AI has traditionally been recommended for avoiding T. foetus but is only practicable in small herds. The replacement of infected bulls with uninfected bulls of less than 3 years of age and culling of non-pregnant cows has been effective in increasing the calving rate of one large infected beef herd5. Given the prevalence of bovine trichomoniasis and the difficulties involved in controlling infection by available means, there is clearly an urgent need for an effective vaccine. Immunization with formalin-fixed whole T. foem and membrane/membrane glycoprotein preparations has been partially successful in the prevention and elimination of infection in bulls39p40.A United States Department of Agriculture conditional licence has been granted to one manufacturer (Franklin Laboratories Inc., Fort Dodge, IA, USA) of a ‘whole-cell’ vaccine. Data, supplied by the manufacturer, showed an apparent 90.6% reduction in infection in ten vaccinated heifers compared with ten control heifers. A preliminary evaluation of a vaccine of unspecified composition from a second manufacturer showed statistically significant differences in infection rates, duration of infection and the firstservice conception rate, between 36 vaccinated and 15 control heifers41. However, the essential question of whether vaccination increases pregnancy rates in the face of natural exposure remains unanswered. In common with other parasitic diseases, the development of a truly effective vaccine is dependent upon obtaining a much deeper understanding of the immunobiology of T. foetus infection. The antigenic dissection of this parasite by monoclonal antibodies is underway and progress has been made in identifying antibodies with demonstrable effector
no.
12. I989
functions such as agglutination, complement-mediated lysis and opsonization by bovine monocytes; one is known that recognizes a surface antigen of approximately 150 kDa42. Two antigens (a 190 kDa protein and a glycoprotein of variable molecular mass) defined by monoclonal antibodies with in vitro effector functions are also being assessed as potential vaccinogens (L.B. Corbeil et al., unpublished) . References 1 Moraan. B.B. (1946) Bovine Trichomoniasis (revised edn) Burgess Publishing Company 2 Dennett, D.P. etal. (1974)Aust. Vet.J. 50,427-431 3 McCoo1,C.J. etal. (1988)Aust. Vet.J. 65,153-156 4 Pefanis, S.M. et al. (1988) J. Afi. Vet. Assoc. 59, 135-138 5 Clark, B.L. etal. (1974)Aust. Vet.3.50,424-426 6 Christensen, H.R., Clark, B.L. and Parsonson, I.M. (1977)Alcst. Vet.J.50,132-134 7 Kimsey, P.B. et al. (1980) 3. Am. Vet. Med. Assoc. 177,616-619 8 Skirrow, S.Z. et al. (1985) 3. Am. Vet. Med. Assoc. 187,405407 9 Goodger, W. J. and Skirrow, S.Z. (1986)J. Am. Vet. Med. Assoc. 189,772-776 10 Clark, B.L., Dufty, J.H. and Parsonson, I.M. (1983) Au-t. Vet.J.60,71-74 11 Rae, D.O. (1989) 3. Am. Vet. Med. Assoc. 194, 771-775 12 Clark, B.L. (1971) in Veterinaty Review No. 1 pp S-26, University of Sydney 13 Roberts, S. J. (1986) in Veterinary Obstetrics and Genital Diseases p. 447, Woodstock 14 Parsonson, I.M., Clark, B.L. and Dufty, J. (1974) Aust. Vet.J.50,421-423 15 Parsonson, I.M., Clark, B.L. and Dufty, J. (1976)J. Coma. Pathol. 86,59-66 16 Abbitt, B. (1980) in Current Tkapy in Theriogenology (1st edn) (Morrow, D.A., ed.), pp 482-488, W.B. Saunders 17 Ball, L., Dargatz, D. and Mortimer, R.G. (1987) in Vetetinary Clinics of North America: Bovine Reproduction (Gibbs, H.C., Herd, R.P. and Murrell, K.D., eds), pp 561-577, W.B. Saunders 18 Skirrow, S.Z. and BonDurant, R.H. Am. 3. Vet. Res. (in press) 19 Bartlett, D.E. and Hammond, D.M. (1945) Am. 3. Vet. Res. 6,91-95 20 Robertson, M. (1963) in Immunity to Protozoa (Garnham, P.C.C., Pierce, A.E. and Roitt, I., eds), pp 336-345, Blackwell Scientific Publications 21 Skirrow, S.Z. (1987) 7. Am. Vet. Med. Assoc. 191, 553-554 1 22 BonDurant, R.H. (1985) Cumpendium of Continuing Education (jar the Practicing Veterinarian) 7, S179-S188) 23 Rhyan, J.C., Stackhouse, L.L. and Quinn, W. J. (1988) Vet. Pathol. 25,350-355 24 Fitzgerald, P.R. (1986) in Veterinary Clinics of North America: Food Animal Practice, Vol. 2 (BonDurant, R.H., ed.) pp 277-282, W .B. Saunders 25 Clark, I.A. and Chat&i, G. (1988) Am. J. Trop. Med. Hyg. 39,246-249 26 Honigberg, B.M. (1970) in Immunity to Parasitic Animals (Jackson, G. J., Herman, R. and Singer, I., eds), pp 469-550, Appleton-Century-Crofts 27 Skirrow, S.Z. -&d BonDurant, R.H. Am. J. Vet. Res. (tn press) 28 Ito, Y. et al. (1975)Jpn.J. Parasitol. 24,333-339 29 Diamond, L.S. (1983) in In vitro Cultivation ofProtozoan Pa&es (jar&, J.B., ed.), pp 89-G, CRC Press 30 Abbitt, B. and Ball, L. (1978) Theriogenology 9, 267-270 ”
I
Parasitology Today, vol. 5, no. 12, I989
31 Tedesco,
32 33 34 35 36 37
L.F., Errico, F. and Del Baglavi, L.P. (1979)Aust. Vet.J.55,322-324 Yule, A. etal. (1989) Vet. Pa:rasitol. 31,115-123 Fitzgerald, P.R., Johnson., A.E. and Hammond, D.M. (1963)J. Am. Vet. Med. Assoc. 143,259-262 McLaughlin, D.K. (1968)J. Purusitol. 55,1038-1039 Williams, O.J. etal. (1987)Awt. Vet.J.64,15%160 Retief, G.P. (1978)s. Afr. Vet. Clin. 23 Miller, M. (1982) in Nitroimiduzoles (Breccia, A., Cavalleri, B. and Adams, G.E., eds), pp 133-148, Plenum
377
38 Palladino, M.R., Campero, C.M. and Villar, J.A. (1982)Gac. Vet. 44,165-174 39 Clark, B.L., Dufty, J.H. and Parsonson, I.M. (1983) Aust. Vet.J.60,178-179
40 Clark, B.L., Emery, D.L. and Dufty, J.H. (1984) Ausr. Vet.?. 61,65-66 41 Mortimer, R. (1988) in Proceedings of the Western Conference for Food Animal
Disease Research
p.6
(Abstr.) 42 Burgess, D.E. (1986) Exp. Parasitol. 62,266-274
Dormancy and Hatching of Nematode Eggs R.N. Perry The sequence of events during the hatching process of some parasitic nematodes has been the subject of recent detailed study. Certain factors, such as the trehalose content of the perivitelline fluid and the permeability characteristics of the eggshell, are influential in hatching and are also importantfor the survival of dormant, unhatched juveniles. In this review, Roland Perry examines some attnbutes of the egg that are involved in dormancy and survival of nematodesand detailsthe changes thatoccur during the hatchingprocess.
Recent reviews of the hatching mechanisms of nematodes’>2, and specifically of the plant parasitic cyst nematodes3, have demonstrated variations in hatching mechanisms which obviate consideration of a detailed ‘model’ hatching system. However, there are certain features of the nematode egg that pertain to hatching and survival in many species. This review concentrates on these features using selected nematode species to illustrate the link between dormancy and hatching. The life cycle of animal and plant parasitic nematodes is frequently synchronized closely with that of their hosts to optimize the chances of successful invasion. This synchrony is often centred on the ability of the unhatched infective juvenile to remain dormant until suitable conditions for hatching occur, and the stimulus for hatching may be provided by the host itself. In some: animal parasitic nematodes this is achieved after the egg has been ingested, with the host gut providing the conditions for hatching. In some plant parasitic nematodes, principally the cyst nematodes,, hatching occurs in response to diffusates from the host roots. With other species that hatch freely in water, root diffusates enhance the rate of hatching to ensure that a large number of infective juveniles emerge close to the growing roots. Thus, the: ability to survive in a dormant state and. the subsequent hatching of infective individuals are interrelated and both may require the same physiological, behavioural and morphological adaptations by the parasite in order 0 1989, Elsev~er Science PubIshers Ltd. (UK) 0 I694707/891802 03
to exploit small, discontinuous host environments. Several of these adaptations have been the subject of recent research, prompted primarily by the need for novel control strategies based on disruption of the nematode’s life cycle. Even among species that depend on a stimulus from the host to induce hatching, the sequence of events from reception of the stimulus to eclosion varies considerably. However, one feature that is involved in hatching mechanisms of several species may also be important to dormancy and survival, namely, the presence of trehalose in the perivitelline fluid surrounding the unhatched juvenile. Importance of trehalose Eggs of Ascaris suum remain viable in
the soil for several years, but ingestion by the host and exposure to the stimulus provided by the alimentary tract causes hatching. Clarke and Perry4 proposed that hatching is initiated when carbon dioxide gas penetrates the lipid layer and becomes hydrated, changing from a linear, relatively non-polar molecule into the larger disc-shaped carbonic acid and its ions; a mutual repulsion between the ions and the ascarosides of the lipid layer results in a structural change in the lipid layer, which alters from a largely impermeable structure to a permeable one with sufficiently large pores to allow the passage of trehalose and enzymes. This change in permeability has been detected by the release of trehalose into the surrounding medium’, thus gradually reducing the
Entomology and Nematology Department
,L+RCInstitute of&-able Crops Research Rothamsted Experimental Station
Harpenden UK
HertfordshireAL5 2JQ,
