EXAGEN – Why Start from Day 1?

EXAGENWhy Start from Day 1?

Decrease the Risk of Crypto with the 3 day Preventative Plan – In Advance of Treatment – just like Vaccination Protection

8 August 2017

Introduction
By referring to the review of work published in Australia (Gunn et al 2016),  we get an insight into just how many areas exist where management has an impact on how crypto infects a calf shed. Faubert & Litvinsky (2000) reported results without the sensitivity of the advanced techniques used by Gunn et al (2016) to identify pathogens. Faubert & Litvinsky only examined cryptosporidial risk and not all pathogens.

Reference Gunn et al (2016):
Note: One farm may not represent all farms. It does represent the infective agents identified day-by-day during the first 3 weeks of calves in a calf shed.
This work showed that on this farm, rotavirus expressed itself earlier in the calfpen environment. It has a shorter prepatent period. It may also show that it is more difficult for farmers to manage once it is established. Other data published as graphs identify the features of the shed which show the increase in cryptosporidium with time.

How does this preventative role for Exagen compare with a vaccination Program?
The role vaccination plays is clear – decrease the severity of signs, improve the rate of recovery, and decrease the virus presence before the shed is loaded with (rota) virus – one of prevention before it gets established. These are exactly the same goals for EXAGEN against Cryptosporidium parvum.
EXAGEN from Day 1 – a different type of a preventative plan – Needs to start before 3 days of age if it is to be compared with a vaccination plan, similarly preventing the take off/ sudden escalation of infection.
There has been much speculation of the possibility of a vaccine for control of cryptosporidiosis. So far it has not been successful, and to the best of my knowledge no recent progress has shown either antibody and cellular immunity can be developed and be available for when the protection is required. The closest farmers can go for crypto scours is to use Exagen on the farm with a history of cryptoscours – not much different from rotavirus vaccination really, relying on a different way to mitigate the risks of the respective diseases. Get in early before crypto is expressed and before it is spread between calves.
By examining the graphs in that review and concentrating on crypto scour specifically, the calves are excreting oocysts well in advance of apparent significant contamination of pen walls, and calf pen floors. It would be reasonable to say we do not know exactly how the infection appeared in 3 -5 day old calves. We do know that there is a risk of crypto infection from the calf’s mother either in the calving down process or early in the calving down area. However Gunn work shows that the tractor tray is a real risk factor. It suggests that it requires rigorously cleaning, repeatedly, probably every day or second within the same season (Gunn et al 2016). It is known that an incubation period of 3 days would be required before these infections appear in a calf (Thomson 2016).

Is Halocur® an alternative?
It is important that calves which do start to excrete around day 3 are prevented from developing clinical scours, and that this is not just a delaying of the excretion of oocysts (as for Halocur treated calves) but that oocysts are destroyed so that any multiplication intra-intestinally is halted. This is Exagen’ target – death to oocysts while multiplying in the gut, not just putting up a stop sign and then letting them go – 7 days later to infect the shed and contaminate the handlers and the general environment.

Castro-Hermida et al (2003) research shows that by giving beta-cyclodextrin 3 times daily to mice it is possible to prevent 99.4% of infections becoming established. This compares with 70% with twice daily treatment with betacyclodextrin. The total dose per day is the same 25 g in either 2x (12.5 g) or 3x (8 g) split doses. Three days of treatment are required to knock out infections in mice and stop the re-infection cycle from recommencing. This activity would be included as part of the mitigation action plan described by Moore and colleagues from the Washington State University (2012). Other parts of the plan include hygiene protocols for daily cleaning, and biosecurity protocols to reduce potential spread within the unit.
Without Exagen it is difficult to plan, when the calves remain in an environment slowly becoming overwhelmed by excretion from calves with clinical signs.
Calf to calf spread can only be prevented if calves stop excretion – a task that Exagen is ideally suited (Castro-Hermida et al, 2001). There is no surge from the re-expression of inhibited stages as happens with Halocur (Silverlas et al. 2009)

Knock it down once and stop cryptosporidial scours in its path. Go to the source from day 1 or as soon as the calf is isolated from its mother and in the calfpen. If this is longer than 3 days then consider a longer period of Exagen use than just 3 days.

Additional Factors in the Ultimate Calf Gut Protection Plan:
1. Protect and Stimulate the Natural Gut Flora/The Role of Probiotics:
Optimise the gut development, its natural gut flora, decrease stress which upsets the establishment and potential loss of the Lactobacillus spp. and the influence they have as natural beneficial strains.
The difficulty in recommending a specific probiotic is that the commercial availability of these very specific strains is lagging behind the science. In the species that are desirable there are limited strains within it that have the required characteristics. Hence Lactobacillus reuteri (some strains) and Lactobaciullus plantarum (some strains) may be listed on a product but even then not have the profile required.
In general though
• Found naturally from birth.
• Are inhibitory of other like Lactobacillus species . L reuteri (Christiansen et al), and L plantarum (Rodriguez-Palacios et al, 2017) may have the required profile.
• Some strains of these species are inhibitory of bacterial species such as E coli, and salmonella amongst others. (Rodriguez et al, 2009; Ratsep et al, 2017)
• They are of bovine origin. Bovine strains are essential if they are to be retained to become established.
• Are acid resistant to pass through the stomach.
• Are temperature robust for storage and transport.
• Deliver the correct number of bacteria – generally more than 1 x 109
• Define if there are prebiotics that promote the probiotics strains targetted.

In general the Lactobacilli strains identified are slower to establish themselves as important species in the developing gut, and they are sensitive to stresses (Fuller 1989) such as transport, especially moving up and down ramps, rough handling or surgical interventions – castration or dehorning or changes to feed patterns. The beneficial Lactobacilli spp take time to overcome ‘weedy’ species such as E coli that readily invade from soon after birth.

2. Colostrum’s Role in Gut Development:
Colostrum is important for maturing the different parts of the intestinal tract (Buhler et al, 1997). It works in tandem with the gut microbiome by supplying hormonal and growth factors acting throughout the gut. This maturation helps prevent the adhesion of pathogens, and advances nutritional digestion and absorption of food at the same time (Hammon & Blum 1997).

Summary
a) Hygiene – recognise risk factors and keep to good housekeeping (biosecurity). Refer to Disinfection Part I, Part II
b) Exagen from Day 1 to mitigate the output and cross-contamination of the new calves. Use it for three days for a clean shed, or 7 – 10 days for a shed showing calf scours.
c) Manage stress. Gentle management, develop and maintain routines in the shed.
d) Continue colostrum for its growth factors and not just its antibody content. These growth factors are present in early lactation phases of the cow.
e) Probiotics but you may need some assistance to decide which species and strains within species are more effective.

References:
Buhler C, H Hammon, GL Rossi, JW Blum (1997), Small intestinal Morphology in Eight-Day-Old Calves fed Colostrum for different Jurations or Only Milk replacer and Treated with Long-R3-Insulin-Like-Growth factor I and Growth Hormone Amer Soc of Anim Sci 76: 758-765
Castro-Hermida J A, Y Gonzalez-Losada, F Freire-Santos, A M Oteiza-Lopez, A Ares-Mazas Unexpected activity of b-cyclodextrin against experimental infection by Cryptosporidium parvum. J Parasit (2000), 86 1118-1120
Castro-Hermida J A, G L Yolanda, F Freire-Santos, M Mezo-Mendendez, E Ares-Mazas. Evaluation of b-cyclodextrin against natural infections of cryptosporidiosis in calves. Veterinary Parasitology 101 (2001) 85-89
Faubert GM, Y Litvinsky, 2000. Natural Transmission of Cryptosporidium parvum between daks and calves oin a dairy farm. Parasitology 86 495-500
Fuller R. (1989). A Review: Probiotics in man and animals.Journal of Applied Bacteriology. 66, 365-378
Gunn A, J House (2005). Calf Scours in Southern Australia. Beef Enterprises Phase 2. Project code: AHW.057 Published by Meat & Livestock and University of Sydney. Australia Gunn A, J House, P Sheehy, A Thompson, D Finlaison, P Kirkland (2016), B.AHE.0025 Molecular methods for detection of calf scour pathogens. Pub.: Meat and Livestock Australia Limited The University of Sydney and NSW Department of Industry and Investment. Hammon H,JW Blum (1997) Prolonged Colostrum feeding Enhances Xylose Adbsorption in Neo-natal calves. J Anim Sci 75 2915-2919
Lee Y-K, KY Puong, AC Ouwehand (2003). Displacement of bacterial pathogen from mucus and Caco-2 cell surface by lactobacilli. Jl Medical Microbiology 52, 925-930
Moore AD, K Heaton, S Poisson, WM Sischo (2012) Dairy Calf Housing and Environment: The science Behind Housing and On-Farm Assessments. Pub Washington State University Extension: EMO 045E
Ratsep M,S Koljaig, E Sepp, J Scmidt et al (2017) A combination of probiotic and prebiotic product can prevent the germination of Clostridium difficile spores and infection. Anaerobe 47 94-103
Rodriguez-Palacios A, HR Staempfli, JS Weese (2017) High doses of Halotolerant Gut-Indigenous Lactobacillus plantarum reduce Cultivable Lactobacilli in Newborn Calves without Increasing Its Species Abundance. International Jl of Microbiology, 2017, Article ID 2439025, 11 pages
Rodriguez-Palacios A, HR Staempfli, T Duffield, JS Weese (2009). Isolation of bovine intestinal Lactobacillus plantarum and Pediococcus acidilactici with inhibitory activity against Escherichia coli 0157, and F5. Jl of Appl Microbiol 106 393-401
Silverlas C, C Bjorkman, A Egenvall: (2009). Systematic review and meta-analyses of the effects of halofuginone against calf cryptosporidiosis. Preventive Veterinary Medicine Jl. Prevent med.2009.05.003
Thomson, Sarah (2016) Cryptosporidiosis in farm livestock.PhD thesis. Submitted in fulfilment of the requirements for the Degree of Doctor of Philosophy, Institute of Biodiversity Animal Health and Comparative Medicine University of Glasgow 2016.

Halocur® is the registered trademark of MSD Animal Health.
Copyright: Bruce Pauling B.V.Sc.
8 August 2017

Risk Factors: Calf Scours

Cryptosporidiosis Risk factors: Scientific report on Calf Shed Infections
4 August 2017

Introduction: This report from Australia advances our understanding of spread of rotavirus and cryptosporidiosis as well as other calf scour pathogens using advanced molecular methods.

Gunn et al (2016) report on two farms with calf scour pathogens. She identifies the sequence of events surrounding two sets of infections and monitoring with sensitive  test methods for important calf scour microrganisms. In an earlier report by Gunn (2005) developed an extensive list of risk factors suitable for management. In this latest report a research team goes further and advances our understanding of which of those risk factors may be influencing cases of clinical calf scours. This latest work  reports use new technological methods, describe their development, aimed at  advancing the reliability and understanding of the epidemiology of the important causes of calf scours; these being in particular rotavirus, and  cryptosporidiosis. In this report (Gunn et al 2016) discusses the scientific methods in detail. The results of applying the work included field examples of  two ‘problem’ calf sheds to advance the understanding of how ‘breakouts of infection’ occur inspite of farmers applying recognised standards of care and hygiene. The techniques developed and used to demonstrate more precisely the origins of infection on farms includes the use of swabs to test for rotavirus and cryptosporidiosis on a number of surfaces including calf shed walls, gates, tractor trailers, and calf feeders including feeding teats. And also included calf excretion of the agents being studied. These tests were monitored over time; generally over 3 weeks to reach some specific conclusions for each of the two farms.

One aspect of the calf scours that applied to both farms was the importance of mixed infections. And while acknowledging the presence of both infections (rotavirus TypeA) and cryptosporidiosis, the scope of the research may have limited the interpretation of how important, or relevant to the results such combined infections may have had on the clinical signs seen.  In unpublished work, David & Millward (2005)   report that mixed rotavirus and cryptosporidial infections complicates the interpretation of the time sequence from infection; the appearance and severity of signs are delayed when rotavirus and cryptosporidiosis are infected simultaneously compared with each agent alone.  An important variable not available for Gunn et al ‘s report (2016) is new research on the importance of  the genotypic strain of Cryptosporidium parvum as reported by (Thompson 2016) on the severity of clinical signs. While this is not proven in her PhD thesis the results leave open that cryptosporidial subtypes or strains differ significantly in virulence, and how pathogenic different genotypic strains may appear from a clinical perspective, notably how severe the scours (tendency to dehydrate), the loss of appetite,  and period of recovery from scour can differ. In the practical farm situation there is a lack of knowledge about cryptosporidial challenge dose rate occurring, the inability to measure ‘stress’ in the very young calf, and in general the imprecise measurement of colostral transfer while following high standards of colostrum feeding to a group of calves.  Taking these variables into account it is therefore difficult to apply the results above (Gunn et al 2016) to any other farm other than the two reported on. This report however does emphasise the presence of both organisms, one a virus and the other protozoan and their widespread presence in the calf rearing units from birth and in the shed environment. The use of these new tools to detect their presence is an advance in our understanding. They emphasise the importance of between calf transmission in advance of clinical signs of scouring.

Results:

Both rotavirus and Cryptosporidium parvum were found on a number of surfaces after swabbing.

There were some differences between farms and the report discusses these in further detail.

The use of Halocur on one farm (B) delayed significant shedding of oocysts,  with only 30% shedding before 14 days. Halofuginone is considered to be more effective with lower challenge doses.

These reports produced on these two farms are open to a number of opportunities to propose what and how these infections occurred. These comments (BAPauling) do not necessarily reflect the conclusions of the report prepared by Gunn et al in her report. The full report should be examined to become familiar with each farm’s management and the interpretation of the results obtained.

Details of the findings are attached. The comments are made from a personal perspective and should not be seen as criticism of the management of the calf units or the research undertaken and reported on.

 

Hygiene standards in preparation failed to deliver a clean pen wall. “Hardy” viruses (includes rotavirus) and surface disinfection requires sterilisation standards to be achieved to reduce risks. Multiple cleaning attempts e.g steam cleaning repeatedly and allowing to dry (pers comm.). Sterilisation of the surface means a smooth wall structure – non-porous, with no residual organic matter.
This will not address the problem of the carrier cow excretor infecting the calves.
Interpretation, the floor bedding is clean at the start,
with the contamination subsequent to the expression of infection from wall to calf so that there is rotavirus excretion to floor (day 3 – 5),
Or
Contamination from calf infection from the dam (and fomites) at birth and a new infection expressing itself as contamination over the bedding. The graph below suggests that 40% of calves are excreting rotavirus from about 3 days and it is the dam and/or fomites including feeding utensils which is important.
Once rotavirus infects one part of the system, the wall, or from the cow, the virus becomes ubiquitous – everywhere and transfers to feeding equipment, meaning there is virtually no rescue available to prevent further infection.
The goal without hygiene practices changing may be limited in scope and success.
In the season reductions in severity may be possible – reducing the length of time of scouring may be the best goal.
It is reported that empty pens were cleaned with Virkon, a product which is known does not inactivate cryptosporidial oocysts. A change of surface preparation is required, using physical and then chemical disinfection. Very low levels (e.g 10 – 60 oocysts) may have been enough for infection to start from walls and recycling of crypto in a calf
Refer to Practical Disinfection (www.pvd.co.nz)
It is possible that the rotavirus mixed infection was delaying the expression and recycling in the calf with mixed infections. This appears to be a likely explanation. (David & Millward 2005).
The graph suggests that less than optimal hygiene standards of feeding equipment reflected by crypto presence associated with feeding equipment before and around day 9 and with the prepatent period of 2 -3 days, this difference is the prepatent period for cryptosporidial infections. i.e. bedding contamination occurs at days 12-14.
What is important is that a significant number of calves are excreting prior to contaminatiomn of feeding utensils. So earlier transfer between calves may mean cow to calf, calving-down pad transfer, or transfer from trailer contamination.
See comments above.
Detection of crypto on feeding equipment is compounded with the number of calves excreting in the first 9-11 days. The origin of this expression at day 9-11 on feeding equipment of the expression suggests direct contact cow to calf, and calf to calf may be more important for crypto to spread the problem within the calf pen environment in the first 7-10 days. The mixed infections with rotavirus delayed expression of signs of crypto.
Competitive exclusion of crypto expression when mixed infections, but generally more serious clinical signs occur than that of crypto infection alone (David & Millward).
The important feature is the detection of crypto before 9-11 days. It was only then that there was detection on penwalls, and feeding equipment, followed by bedding. See comments above.
Infection with rotavirus was simultaneous with that of crypto, but with a shorter prepatent period the infection of rotavirus appears to pre-exist that of crypto. This may not truly reflect the source of the calf shed infections; cow to calf, calf trailer to calf while the calf is in transit from calving down to the calf shed.
If rotavirus is present the infectivity rate and % of calves infected reaches a high level of the calves at risk.
If crypto is present with only a low number exposed at the start of life, ultimately the infection rate will reach 100% in the period prior to 3 weeks. The shift from infections to clinical scouring sick calves may be more related to the genotypic strain, load dose ingested of oocysts (Thompson 2016 ), colostrum feeding, gut microbiome, stressors and availability of attachment sites in the immature gut lining. Included in this gut manageement is the use of probiotics, though there is no consensus as to which strains may offer ‘real’ and meaningful benefits.

Conclusions:
1. Mixed infections of young calves less than 3 weeks, of rotavirus and cryptosporidiosis started probably from the cow or the calving environment/trailer (or the transfer from fomites) in the first few hours/days; very early in the calf’s life.
2. The multiplication and shorter prepatent period for rotavirus overwhelms the appearance of cryptosporidiosis infection.
3. A faecal sample of early aged calves (less than 6 – 8 days) in mixed infections may therefore miss many of the pending infections of crypto.

Disinfection and Hygiene:
In my opinion there requires to be a change to disinfection materials and methods in the preparation of calf pens. And without more rigorous hygiene protocols for cleaning of feeding utensil any attempt to see the effects of change to disinfection procedures is likely to fail. The finding that the organisms can be found on trailers used to transfer calves also identifies more rigorous standards eg regular water blasting and steam cleaning. the same advice is provided by Moore et al (2010, 2012).

References:
David & Millward. Compositions and vaccines containing antigen(s) of Cryptosporidium parvum and of another pathogen. USA Patent 2005 US 20050106163 A1
Gunn A, J House (2005). Calf Scours in Southern Australia. Beef Enterprises Phase 2. Project code: AHW.057 Published by Meat & Livestock and University of Sydney, Australia
Gunn A, J House, P Sheehy, A Thompson, D Finlaison, P Kirkland (2016), B.AHE.0025 Molecular methods for detection of calf scour pathogens. Pub.: Meat and Livestock Australia Limited The University of Sydney and NSW Department of Industry and Investment.
Moore DA, K Heaton, S Poisson, WM Sischo (2010). Calf Housing and Environments Series. V. Reducing Pathogen Load in the Calf Environment. Pub Washington State University Dec 2010
Moore DA, K Heaton, S Poisson, WM Sischo (2012). Dairy Calf Housing and Environment: The Science Behind Housing and On-Farm Assessments. Pub. Washington State University
Thompson, Sarah (2016). Cryptosporidiosis in farm livestock. PhD thesis. http://theses.gla.ac.uk/7096/. Submitted in fulfilment of the requirements for the Degree of Doctor of Philosophy, Institute of Biodiversity Animal Health and Comparative Medicine. University of Glasgow 2016

July 2017:
Bruce Pauling is a veterinarian and director of Professional Veterinary Distributors, having researched, and developed Kryptade and Exagen registered veterinary medicines in New Zealand for aiding the prevention and treatment of calf scours associated with Cryptosporidium parvum.

Disinfectants for Practical Cryptosporidium Management

Updated: Information of Disinfectants:  17 July 2017

Introduction:

Numbers of reviews state that Cryptosporidium is difficult to kill. This applies to municipal water treatment for human risk management and the farm shed to protect and prevent clinical cryptosporidiosis for calves. Cryptosporidial oocysts are at least 15 x more resistant than Giardia to disinfectants. In mild infections without scouring, the output of infective stage oocysts is still far above an infective dose. In fact there is still a possibility that non-detectable and inapparent infections are occurring because of the insensitivity of the tests systems. With a scouring calf there are massive numbers of oocysts produced.
Therefore the activity of any active ingredient of a disinfectant requires to be

  • very high in the face of such a challenge when applied for control.
  • ideally it should be safe to use, and
  • environmentally ‘friendly’.

Assessments of Performance

Assessment of effectiveness against the infective oocyst may be made by a number of in vivo, and/or in vitro tests. Different results in performance can arise by using different techniques.

Establishing a relative performance for each ingredient, leads to studies on

  • each concentration recommended,
  • a comparison of the time of contact for the relevant concentration.
Brand names Ref Active ingredient Availability Comments
Neopredisan 2-4% Keidel & Daugschies 2013 p-chloro-m cresol Not available in New Zealand
KenoTMCox 2-3% Naciri et al 2011 Amine based formulation Not available in New Zealand
Ox-Virin 10% Quilez et al 2005 25% hydrogen peroxide, 5% peracetic acid An alternative brand formulated with the same active ingredients is available in New Zealand Dangerous goods with strict guidelines on Protective Clothing
Peroxide 3% (No brand) 6% is recommended for higher performance

Comparison of Disinfectants and Methods of Decontamination:

Chemical Disinfection

Against this background there are a number of brands which have been assessed for efficacy in the laboratory using in vivo and in vitro methods. A number of active ingredients have not performed to give any assurance of effective disinfection when faced with the challenge of very high numbers of oocysts in a calf shed.
One independent group which has checked research and reached some conclusions is Moredun Research Institute in Scotland (Hotchkiss et al 2015). Their nominated brands do not relate to what is available in New Zealand, but it does emphasise how difficult it is to transfer benchtesting to actual live field performances in the face of an outbreak.
None of the products above are recommended be used in a shed with livestock, but they are used for shed disinfection. All require protective clothing including masks.
Others options are based on ammonia and hydrogen peroxide provided they are used at the correct concentration and given the appropriate contact time. Safety measures will be required for these products as well.

Physical methods:
  1. Steam cleaning is safe and effective.
  2. Decontamination with water blasting can be an important first step in disinfection, but in a calf shed this is not suitable if the bedding becomes wet. It may be suitable for grated or concreted areas on a regular basis e.g. daily or every second day when calves are present. Though mists created by high pressure hosing may create a problem of vaporisation and spread of oocysts over the surrounding areas.
  3. Drying of calf faeces (spread out thinly) for over 3-4 days is effective in preventing oocyst survival (Anderson 1986).
  4. Temperature effects are overridden by moisture, moisture being protective esp with colder temperatures, with months recorded of survival in damp and cool conditions. Warmer temperatures over 15 degrees reduces infectivity to over 2 weeks, or faster with higher temperatures.
Other (NZ) active Ingredients
Brand names Supplier Claim
Vetsan Vetpak “Stabilised” chlorine dioxide Amount used and contact time varies. Has recommendations for use in calf shed. Stabilised activity claims has yet to be measured against the gold standard of mice infectivity testing.
Swift Ethical Agents Chlorine dioxide (2 pack) Must be used fresh, and is extremely pungent. Unable to be used in shed with calves present Traditional source of chlorine dioxide.

Other active ingredients such as chlorine dioxide are sometimes recommended than those above but these fail to be listed as preferred products. There are varying opinions on their usefulness.

Below is a summary of research reported in the literature:
Chauret et al 2001 reports for example on tests completed on purified oocysts originating from three different suppliers. Results showed marked differences with respect to their resistance to inactivation when using chlorine dioxide. Contact time values of 75, 550, and 1,000 mg . min/liter were required to achieve approximately 2.0 log10 units (lowering by a hundredth) of inactivation with oocysts from different sources.

References in the literature often refer to them performing well in the face of lower numbers of challenge oocysts. Their performance when there are higher levels of oocysts challenging doses as found in a contaminated calf shed with endemic cryptosporidial scours may not be the same however.

Liyanage et al., (1997) determined that it is the un-dissociated chlorine dioxide that is the main anticryptosporidial component. The component parts do not have any significant effect, against the cryptosporidial oocyst e.g. chlorite, chlorates or chlorides. Given the instability of the chlorine dioxide and the narrow pH at which it is present/active, the speed at which it becomes inactivated, claims of stability need to be substantiated in a critical manner, using the gold standard of mice infectivity studies, and at varying concentrations and over variable contact times with varying number of oocysts of known infectivity in mice.
One report on chlorine dioxide by Chauret (2001) summarises a number of trials (11) and assesses the contact time, the degree of inactivation, and the test method used. It shows that for a lowering by a hundredth or a thousand, a contact time of more than 2 – 6 hours was generally required especially when the mouse infectivity gold standard was used. Cell culture tests used as an alternative test method demonstrated a wider variety of contact times. Note: it was not possible to lower oocysts to a useful degree of over 1000 fold reduction, which is the useful measure of disinfection for cryptosporidial oocyst control. A threshold of 99.5 % inactivation is proposed to evaluate disinfectants in vitro using C. parvum as model organism (Dresely et al, 2014). On this basis chlorine dioxide would fail to be classed as a disinfectant.
Trial work takes into account a large amount of information and for each comparison, the same conditions may not be able to be compared with a second trial result. For completeness the comparisons made include the following as reported by Casemore & Watson

Physical Variables Chemical Variables Study Design Variables
Temperature during test exposure Disinfectant type and test concentration; method of measuring concentration (e.g. start, end, integrated) Oocyst source and strain characterisation
Test system pH Disinfectant neutralisation prior to viability/infectivity testing Oocyst clean up method, age and storage conditions.
Contact time Composition of disinfectant (e.g. presence of surfactants or other agents in test product) Oocyst test concentration used; accuracy of counts
Effect of physical degradation and environmental stress on oocysts. Denaturing/dissociation of disinfectant during contact period; method of measuring Method of assessing efficacy – in vitro (excystation, sporozoite ratio, cell culture), infectivity in vivo.
Nature of test medium and organic load*. Method and timing of measuring disinfectant concentration (batch vs continuous, etc).

Conclusions made by such summaries include

*Size of the oocyst inoculum: the CT (Contact Time) value may be greatly affected by the number of oocysts present at the beginning of exposure to a disinfectant. This appears to have been orders of magnitude higher in some studies, e.g. ozone was reported to be the least effective of all the studies. Numbers of oocysts present may markedly affect the dynamics of disinfection.

Measuring Oocysts Viability and Infectivity

Background:

The load of oocysts is highly variable in an infected calf shed. Identifying how much ‘risk’ actually present is difficult.
The oocysts identified as present also vary in viability. The number required to infect and lead to clinical scours also varies with genotype strain and it also changes as oocysts age and lose infectivity. This is partly from their exposure to the environment. Temperature and moisture are critical conditions determining survivability and infectivity. Faeces dried to the air require only 4 days to be become non-viable. Wet and colder conditions lead to extended periods of infectivity of over 6 months.
The gold standard to assessment of viability is infecting newborn mice and measuring infection intensity in the intestine.
In vitro (without using mice) have also been reported and have been compared with mice infectivity studies. In vitro tests underestimates the loss from disinfection procedures when compared with the gold standard (mice testing).
Because numbers of oocysts in the calf shed are going to be extremely high – the efficacy of any disinfection needs to be at an extremely high level of over 99.9%, otherwise there will be sufficient levels of oocysts left to provide an infective dose.
Expectations that this can be achieved while calves are in the shed are unrealistic with the known chemicals available that can achieve this efficacy.

Young calves are highly susceptible to becoming infected and expressing the clinical signs of scouring with loss of appetite. This is emphasised when considering the fact that some young calves get infected with as few as 10 oocysts (50% of a group of calves: 60 oocysts). The level at which cows are excreting is at a level which is lower than the sensitivity of the commercial laboratories to detect oocysts. Less orthodox test protocols are required to concentrate the oocysts in adult cow pats (dung) before counting them.
Part of the variability in the level of oocyst infection required to infect a calf is thought to be related to the genotypic strains that exists between Cryptosporidium parvum isolates.
The very young calf is a very sensitivity indicator that oocysts are present, being infected with doses that are lower than the sensitivity of the tests used to measure the presence of oocysts.

Doing some numbers:

This is especially relevant when assessing how effective a disinfectant or hygiene programme is. In the calf infection with scouring, their multiplication in the gut leads to massive numbers and output of greater than 1 x 106/g (and higher) of dung. The level of output in the cow at parturition with a partial post-parturient rise is about 400-500 oocysts per gram of dung. This is still below the sensitivity of the test (microscopic examination) which is about 1000 oocysts per gm of dung. Other tests can go as low as 100 oocysts (using immunofluorescent methods).  The calf in theory requires about 1/7th g of its dam’s dung to become infected. Detecting the excreting cow is currrently not available commercially.

Conclusions

  • It is unrealistic to expect disinfection to lower the level of infective oocysts in an infected calf shed. The transition shed may be the most suitable for change.

a) Physical removal of dung requires concrete surfaces to be sprayed (physical decontamination) down daily or twice daily, before signs of scours shows.

b) Concentrate efforts in the first 5-10 days especially the first 2 – 3 days in the calf rearing environment.

c) Early detection and separation into a hospital concreted/grated area. Get them when they are depressed and before they scour. Spend time watching for calves separating and becoming isolated. Swiss studies with one strain showed that only about 45% of infected scouring calves will lose their appetite. All of them require to be identified and treated with KRYPTADE for 3 days as on the label.

d.) If there is no transition area for calves EXAGEN is recommended to ensure numbers of oocysts excreted are capped at very low levels. It is not an alternative to low standards of hygiene and cleanliness of feeding utensils. Use EXAGEN twice daily for a minimum of 3 days at the start of the season. If scours becomes established in the calf shed e.g later in the season, this should be increased to up to 10 days or longer.

e) While twice daily split dosing of 25 g with EXAGEN is routinely recommended as optimal, there is another option for higher level control. This is aimed to further decrease cross infection and more effective prevention of new infections by splitting the daily dose into three; i.e EXAGEN three times (5 g for very small calves, or 8 g for larger calves.) eight hours apart if management can be adjusted

  • Because the cow is being recognised as important source of calf infections, the first calf case each season may be an important indicator of cow-calf transmission, keeping good records is aimed to determining which cow may have been responsible for the first infection (and the start of the scouring).
  • Have a broad understanding that disinfection and hygiene standards will include keeping out all visitors/strangers who may carry crypto into your unit. It includes your own risk factors such as clothing and footware as carrying infection/contaminants. If you practice limiting visitors, then enact protocols that address your own risks of transmission within your own unit. Create ‘barriers’ that identify where extra precautions and isolation start applying. Foot baths with scrubbing including footware changes, protective clothing. Clothing and footware spelling 3 – 4 days in sunlight/in warm conditions (winter allowing) before re-using  allows for natural disinfection and reduces any oocysts surviving on footware and clothing. Be critical of your own standards, and be prepared to be strict to ensure that your protocol reducing transmission is being implemented. Just because it doesn’t appear to be practical don’t discard an option. It may be the critical factor making your risks lower.

Watch for developments in cow detection methods to identify carriers.

Be prepared to consider EXAGEN from the first born calf coming into the calf unit especially through the transmission area, to maintain lower levels of oocycst output from inapparent infections. This product is oocystocidal meaning that there is no bounce back excretion of oocysts once treatment is stopped as with halofuginone lactate

Bruce Pauling
4 July 2017

Research Requirements

Research on Cryptosporidium parvum scours in New Zealand.

To better understand the relative importance and to assess risk factors to manage, we need to have more information on the disease and how it is transferred. Can New Zealand contribute to cryptosporidiosis understanding and at the same time develop management protocols to reduce the risk of disease?
Epidemiology: Understanding the climate and management factors affecting between-cow spread of genotypic subtypes of Cryptosporidium parvum has to be one of the high priorities for research.

Genotypic sub-type strain identification: Epidemiology will advance significantly with the information provided with genotype sub-typing by providing a technique to follow infections of selected genotypic subtypes in a population of cows (or calves).

Recent research has indicated that different genotypic subtypes traverse closed (cows) herds during any one year. Not all genotypes are equivalent in severity of signs (Thomson 2016). Early research (Thomson 2016) suggests that the period of excretion of oocysts from one cow may be limited but may be critical with respect to the development of subsequent cryptosporidial calf scours.
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Calf Studies

Research on Cryptosporidiosis in Scotland

Thompson, Sarah (2016) Cryptosporidiosis in farm livestock. PhD thesis. Submitted in fulfilment of the requirements for the Degree of Doctor of Philosophy, Institute of Biodiversity Animal Health and Comparative Medicine University of Glasgow 2016. http://theses.gla.ac.uk/7096/

This extensive and original Scottish research has highlighted some important features of Cryptosporidium parvum infections in calves. Some of the results may or may not have direct relevance to New Zealand farming conditions. In some instances they may be so relevant that they will challenge some of our current practices. Without some local (NZ) studies it is too early to make recommendations.
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