A case of hepatogenous photosensitisation was seen in ewes with lambs at foot grazing an oat crop in the Riverina. While this is the most common type of photosensitisation seen in livestock (Hargis and Ginn, 2007), it is rarely reported on winter cereal crops. The condition results in significant morbidity and represents an animal welfare concern (Smith and Towers, 2002). It leads to stress and reduced immunological competence in affected animals (Smith and Payne, 1991). Economic losses result from associated deaths, condemnation of carcasses, reduced bodyweights (Smith, 2000) and decreased ewe fertility (Morris et al., 1991).
Hepatogenous photosensitisation is a sequel to hepatocellular damage, bile duct obstruction and, less commonly, inherited hepatic defects. The liver is unable to effectively excrete phylloerythrin, which is a breakdown product of chlorophyll formed in the alimentary tract (Hargis and Ginn, 2007). Phylloerythrin is a photosensitising agent and has the molecular structure of a porphyrin. It travels from the systemic circulation into the skin where it absorbs solar energy, particularly in areas that are non-pigmented or exposed, releasing rree radicals and leading to inflammation and tissue necrosis (Barrington, 2014; Tennant and Center, 2008).
In this case, the fungus Pithomyces chartarum was isolated, which is a known cause of hepatogenous photosensitisation. It grows on pasture litter and produces the mycotoxin sporidesmin, which concentrates in bile causing pericholangitis and occlusion of biliary ducts (Smith and Towers, 2002; Waghorn et al., 2002). Alternative aetiologies are also discussed in this paper.
In late June 2014, a mob of 1000 Dohne and Merino ewes with four-month old lambs at foot was put onto a grazing oat crop. Two weeks later 200 of 1000 ewes showed signs of photosensitisation and, of these, 20 were severe. While no lambs became affected, the condition of the ewes deteriorated over the next week and all sheep were subsequently moved off the pasture. A veterinary visit was conducted one week after that and the farmer reported a marked improvement, including a significant reduction in facial oedema, during this time.
The oat crop had been direct drilled into barley stubble three months earlier and had not been treated with chemicals or fertilisers. The producer had grazed sheep on the same paddock sown with oats every year without problems. The only differences this season were increased rainfall and mild autumn temperatures. Additionally, a purple fungus was observed on the barley stubble when the oat crop was sown.
A pasture examination revealed a lush dense green oat crop approximately 60 cm high with occasional plants of capeweed (Arctotheca calendula) and Wimmera ryegrass (Lolium rigidum). In a few small areas, the oat plants had turned purple. The underlying barley stubble had a white fungus growing on it in wetter parts of the paddock.
The water source consisted of two dams that were de-silted over the summer. No supplemental grain, hay or other fodder was provided. Prior to moving onto the oat crop, ewes and lambs were grazing a paddock containing oats, lucerne and clover and approximately 15 percent hairy panic (Panicum effusum) for six weeks.
Ewes in the mob were mixed from multiple sources. The farm manager bought around 200-300 replacement ewes every year. Ages ranged from three to nine years, with the majority of the flock between three and six. Cases were seen across all ages and in both Dohne and Merino breeds.
The severely affected ewes were examined. All were bright, alert and responsive, vital signs were within normal limits andthere was no evidence of icterus. They had extensive epidermal necrosis and sloughing of faces, udders and distal limbs. There was only minimal subcutaneous oedema, primarily on faces. A close examination of skin revealed areas with mild to moderate erythema, ulceration, serous exudation and brown crusting. There was significant epidermal regeneration and new skin was pink and soft. The worst affected ewe had lost the tip of the left ear.
Blood was collected from two severely affected ewes and a full blood count and mean biochemical analysis was performed on each.
Sheep 1 had high GGT: 132 U/L (reference range 47-95 U/L), GLDH: 27 U/L (reference range 1-23 U/L), AST: 248 U/L (reference range 59-130 U/L) and CK: 896 U/L (reference range 1-617 U/L). Sheep 2 had high GLDH: 45 U/L, AST 265 U/L and CK: 761 U/L, while GGT: 89 was within the normal range.
GLDH: glutamate dehydrogenase
AST: aspartate aminotransferase
CK: creatine kinase
Biochemistry for both sheep also demonstrated moderate pre-renal azotaemia and mild hypocalcaemia. Additionally they had hypergammaglobulinaemia suggesting chronic antigenic stimulation. Sheep 1 was mildly anaemic, which may have been of greater significant in light of the moderate dehydration.
The pathologist concluded that the findings were consistent with hepatogenous photosensitisation.
Both purple and normal oat plants, along with the underlying barley stubble, were submitted for analysis.
The fungus Pithomyces chartarum was isolated in spore washes of the barley stubble; however numbers were too small to quantify. No spores were found on the oat plants. No other fungal organisms were seen.
All sheep made a full recovery over a period of four weeks. The severely affected ewes were housed in a shearing shed and fed cereal hay for two to three weeks while the rest of the mob was moved to a different pasture with plenty of shade. No animals suffered secondary bacterial or fungal skin infections, flystrike or other complications.
Pithomyces chartarum is predominantly a saprophytic fungus that inhabits decaying plant matter. While commonly associated with ryegrass dominant pastures, it has been isolated on a range of vegetation (Di Menna et. al., 2009). Some strains of P. chartarum produce spores coated with the hepatotoxin sporidesmin, which is the aetiological agent of ‘facial eczema’ or pithomycotoxicosis (Ludeman, 1999). Collin et al. (1998) showed that 89-100 percent of isolates from New South Wales produce sporidesmin.
In this case, P. chartarum was isolated on samples of barley stubble but not on actively growing oat (Avena sativa) plants. This is consistent with its predilection for senescent and dead plant material at the pasture base. Spores were isolated in numbers too low to accurately quantify. Smith (2000) considered that the minimum detectable level is 5000 spores per gram of fresh grass. In one trial to assess responses to low doses of sporidesmin, sheep were fed the toxin at a dose of 0.0042 mg per kg bodyweight (equivalent to 40,000 spores per gram of pasture) for three to 48 days. The study concluded that sporidesmin causes clinical disease at ‘very low daily doses’, and the effects appear to be cumulative. No lower dose treatment groups were assessed in the trial (Smith, 2000).
In light of these findings, the positive identification of P. chartarum on spore washes may be consistent with a diagnosis of pithomycotoxicosis, despite the low yield. Additionally, the quantity of spores may not adequately reflect the true load on the pasture. Spore concentration is known to fluctuate rapidly over time, and can also vary between sites within a pasture, with sheltered areas and warmer aspects generally having higher counts (Di Menna et. al, 2009 and Di Menna and Bailey, 1973). Finally, there may have been degradation of spores during the time between submission of samples to the laboratory and subsequent fungal analysis, which involved a delay of three weeks. The life span of fungal spores at room temperature varies from a few days to many months (Hoekstra, 2002).
The plant pathologist concluded that isolation of P. chartarum was supportive of a diagnosis of pithomycotoxicosis, although the quantity of spores was too low to be definitive. Spore numbers often decline markedly before clinical signs of photosensitisation are observed and consequently ascertaining accurate retrospective counts is not possible (Priest, M pers comms 5 August 2014). Sampling of the pasture took place nine days after the initial veterinary investigation, and five and a half weeks after introduction of ewes and lambs onto the oat crop, allowing ample time for spore concentration to diminish. Additionally, it seems reasonable that the strain involved was sporidesmin-producing given the dominance of these isolates in New South Wales.
Environmental conditions that promote P. chartarum growth include the presence of decaying leaves and litter, relative humidity of 60 percent and ambient temperatures of 18 to 27⁰C, while optimal sporidesmin production requires a relative humidity of 100 percent and a temperature range of 20 to 25⁰C (Lima et al., 2012). The pH range for maximal growth of P. chartarum is broad extending from 4 to 7 (Ludeman, 1999). High risk conditions for pithomycotoxicosis have been defined as two or more episodes of rain greater than a few mm with concomitant, or immediately successive, grass minimum temperatures of 12⁰C or higher for two or more days (Mitchell et al., 1959). Others specify rainfall of 3.8 mm or more, minimum temperatures of 12.2⁰C extending beyond 72 hours and a relative humidity of 98 percent or higher (Mitchell et al., 1959; Peixoto et al., 2011).
Sheep presented with signs of photosensitisation at the end of June 2014, following an autumn that was noted locally for mild temperatures. In fact, the Bureau of Meteorology released a statement reporting an ‘exceptionally prolonged autumn warm spell’ over much of Australia and records were set in Wagga Wagga in May for the highest minimum and maximum temperatures recorded so late in the season. The preceding summer in Wagga Wagga was notably hot and dry. Relative humidity averages ranged from 69 to 88 from March through to June with maximum values between 97 and 99. Above average rainfall was seen throughout autumn (in March, April and June). Outbreaks of facial eczema are associated with rain and mild temperatures following a hot dry period (Di Menna et. al., 2009 and Mitchell et. al., 1959), which is consistent with the weather pattern seen in this case.
The period between ingestion of sporidesmin and development of photosensitisation is typically seven to 24 days (Di Menna et al., 2009). In this case ewes were first observed with photosensitisation two weeks after introduction onto the oat crop. It is possible that they preferentially grazed the underlying barley stubble to address the fibre deficit posed by the lush green oat crop. Their diet would have also represented a concentrated source of chlorophyll, which forms a critical component in the pathological pathway of hepatogenous photosensitisation.
Pithomycotoxicosis begins with absorption of sporidesmin in the upper gastrointestinal tract. The toxin is rapidly absorbed into the systemic circulation and travels to the liver where it concentrates in the bile ducts. It undergoes redox cycling and generates free radicals causing damage to the biliary epithelium. Bile ducts become occluded with inflammatory exudates and necrotic material and biliary excretion halts, preventing normal clearance of phylloerythrin from the portal circulation. Phylloerythrin is formed when microorganisms in the rumen and large intestine break down chlorophyll. When phylloerythrin is not effectively excreted, it enters the peripheral circulation and accumulates in the skin. In areas that are unpigmented and exposed, it interacts with ultraviolet light resulting in oxidative injury and a photosensitisation reaction (Di Menna, 2009; Smith and Towers, 2002; Tennant and Center, 2008).
While moderate to severe photosensitisation was seen in lactating ewes, their four-month old lambs at foot were not clinically affected. This is consistent with clinical trials in which ewes were dosed with 1 mg per kg of sporidesmin with no detectable levels found in milk (Mortimer and Strandbridge, 1968). Conversely, significant reductions in milk yield have been reported in affected ewes (Rammel, 1991). The lambs in this case appeared to be growing normally but were not weighed. Genetic differences in susceptibility to pithomycotoxicosis have been seen with Merinos shown to be relatively more resistant compared with some British breeds (Di Menna et al., 2009). In this case no variation in incidence or severity was seen between Merino and Dohnes or within lines of each breed.
The severely affected ewes were already recovering well at the time of the veterinary examination. They had been housed in a shearing shed for one week with access to cereal hay and water. The initial marked oedema reported by the farmer had almost entirely resolved. Skin of faces, udders and distal limbs showed extensive epidermal necrosis and sloughing but epidermal regeneration was progressing well and new skin appeared viable. The ewes were not clinically icteric and the farmer was unsure whether they had been earlier in the course of the disease. As the ewes were recovering, a post-mortem examination and assessment of liver histology was not undertaken. Liver injury was assessed by clinical pathology.
The liver enzyme γ-glutamyltransferase (GGT) is considered to be a reliable indicator of sporidesmin-induced liver injury (Ramel, 1991). As well as being used in diagnosis, it has also been applied to breeding programs selecting for resistance to pithomycotoxicosis (Di Menna, 2009). One ewe had a mild increase in GGT, while the other had a value within the normal range. The half-life of GGT has not been definitively established in sheep or other domestic animals, although an estimation of three days has been made in dogs and horses (Hoffman et al., 2008). It is presumed that GGT levels peaked prior to the veterinary assessment. A study of Fasciola hepatica in sheep demonstrated elevated GGT 24 days after treatment with a flukicide but levels had declined markedly during this period (Scott et al., 2005). One ewe also presented with mild anaemia, which can be seen with pithomycotoxicosis (Peixoto et al., 2011). The pathologist concluded that the clinical biochemistry was consistent with hepatogenous photosensitisation.
The aeitiology underlying the hepatogenous photosensitisation is uncertain as there is insufficient evidence to definitively implicate sporidesmin. Mckenzie (2012) describes ‘very rare’ poisonings of ruminants on grazing oats (Avena sativa) causing photosensitisation. There is field evidence of toxicity but the toxin is yet to be identified. McKenzie (2015) hypothesises that it may be associated with plants suddenly producing large quantities of chemicals, such as phytoallexins, in response to insects, microbes or environmental stress.
While there have been numerous publications on pithomycotoxicosis, case reports of photosensitisation on grazing oats are rare, and mostly unpublished. In 1931, Schmidt (cited in McKenzie, 2015) described 5 incidents in sheep and goats in Texas, USA, and published an account in the Texas Agricultural Experiment Station Annual Report for that year. Animals showed signs of photosensitisation after spending 10 days or more on green grazing oats. In 1937, Matthews (cited in McKenzie, 2015) reported unpublished data from Schmidt noting the presence of crystals in the gallbladder of affected animals, suggesting possible involvement of steroidal saponins.
An unpublished report from Gibson in 1999 (cited in McKenzie, 2015) described photosensitisation in a single 10-month old male goat grazing oats in Oakey, Queensland in July 1999. While no icterus was seen on post-mortem, liver parameters including bilirubin, aspartate aminotransferase (AST), glutamate dehydrogenase (GLDH) and GGT were elevated. Scanlan (cited in McKenzie, 2015) provided an unpublished record of hepatogenous photosensitisation in sheep near Coonamble, New South Wales, in July 2003. Again no icterus was seen clinically but AST and GGT were increased in two affected sheep. This flock also had access to the potentially photosensitising plant Caltrop (Tribulus terrestris) and a history of grazing the hepatotoxic weed Paterson’s curse (Echium plantagineum).
There were two large outbreaks of photosensitisation in sheep grazing oat and vetch (Vicia spp) crops in Western Australia in unpublished data from Allen in 2007 (cited in McKenzie, 2015). Morbidity reached 90 percent of 2000 lambs. In July 2014, another case of photosensitisation was reported in lambing ewes in Wagga Wagga following two weeks of grazing an oat crop (Ross, A, pers comm 7 July 2014). There have also been a few cases of primary photosensitisation involving grazing oats; however this appears to be a separate syndrome (McKenzie, 2012).
Prior to introduction onto the oat crop, the ewes were on a different paddock containing approximately 15 percent hairy panic (Panicum effusum). This plant is a common cause of hepatogenous photosensitisation in this region. It contains steroidal saponins that induce crystal-associated cholangiohepatopahty. Young ruminants are most susceptible and often need to consume large quantities for a period of months before developing clinical signs (McKenzie, 2012). Cases are typically seen where P. effusum constitutes 50 percent or more of daily dietary intake (Morton, T., 2010). It seems unlikely that adult ewes, grazing a relatively small percentage of the plant, would have developed photosensitisation as a result, particularly as clinical signs did not appear until two weeks after they were moved to a new pasture.
In two weeks, it is reasonable to expect that the liver would have begun regeneration in the absence of further insults (Mannus, cited in Di Menna, 2009). It is possible, however, that exposure to P. effusum rendered the ewes more susceptible to subsequent liver damage, particularly as they faced high metabolic demands associated with late gestation and early lactation.
The ewes in this case showed signs of hepatogenous photosensitisation two weeks after introduction onto a green oat crop, which comprised their entire diet. Isolation of P. chartarum from underlying barley stubble is suggestive of pithomycotoxicosis, and clinical signs, clinical pathology and environmental conditions support this diagnosis. The low quantity of spores, however, is of uncertain significance and is not definitive for pithomycotoxicosis. It is unlikely that the numbers derived from spore washes accurately reflected the true concentration on the pasture, particularly for earlier points in time. Meanwhile, alternative toxic causes such as phytoallexins and other unidentified chemicals cannot be excluded. Further research is needed to better characterise cases of photosensitisation on grazing oats and potentially identify new toxic principles.
Ross McKenzie author of Australia’s Poisonous Plants, Fungi and Cyanobacteria; Tony Ross Veterinary Pathologist at Charles Sturt University Wagga Wagga and Michael Priest Special Plant Pathologist at the Department of Primary Industries Orange.