Martin Wiedmann, Dr. med. vet., PhD
Listeria monocytogenes is a common pathogen that can cause both animal and human infections. While animal infections have been reported in more than 40 different species, animal listeriosis is most commonly reported in farm ruminants (cattle, sheep, and goats). A better understanding of transmission and ecology of this pathogen in farm animals and in farm environments will not only facilitate better control of this animal disease, but may also reduce human foodborne illnesses.
The genus Listeria contains five species, namely L. monocytogenes, L. ivanovii, L. innocua, L. seeligeri, and L. welshimeri. All members of this genus are gram-positive non-spore forming rods.1 While L. monocytogenes causes both human and animal disease, L. ivanovii is predominantly associated with disease (specifically abortions) in sheep. The other Listeria species are considered non-pathogenic. The pathogenic species L. monocytogenes and L. ivanovii are both hemolytic, as is the non-pathogenic L. seeligeri, while L. innocua and L. welshimeri are non-hemolytic. L. monocytogenes is considered a ubiquitous organism, which can be isolated from many different environmental sources (surface water, soil, sewage, plant material etc.). This organism has the ability to grow and survive under a variety of different conditions. L. monocytogenes grows from close to 0°C to 44°C and is thus considered a psychrotolerant organism. All Listeria spp. are heat sensitive and pasteurization effectively kills Listeria.
All Listeria spp. are phenotypically very similar and their differentiation can sometimes be difficult. Commonly used tests to differentiate Listeria spp. include acid production from D-xylose, L-rhamnose, alpha-methyl-D-mannoside, and mannitol as well as hemolysis patterns on blood agar plates. Hemolysis is usually observed on sheep blood plates, but blood from other species can also be used (e.g., horses, cattle). The CAMP test with Rhodococcus equi and Staphylococcus aureus has sometimes been recommended to clarify hemolysis patterns, but may be difficult to interpret, particularly for people without experience in the use of this test. While virulence tests in mice have historically been used for more definitive speciation, routine pathogenicity testing is generally unnecessary. Molecular and genetic methods generally provide reliable and more rapid means for speciation of Listeria isolates. For example, polymerase chain reaction (PCR)-based methods can be used to screen for the presence of the L. monocytogenes hemolysin gene (hlyA), which is unique to this species, or to screen for other L. monocytogenes specific genes.2
The pathogenesis and genetics of L. monocytogenes have been explored extensively. For many decades, L. monocytogenes has been recognized as a model bacterial pathogen that induces T-cell mediated cellular immunity.3 Studies on the cellular pathogenesis of listeriosis showed that L. monocytogenes is a facultative intracellular pathogen, which has a unique ability to use host cell proteins to spread from cell-to-cell. The ability of L. monocytogenes to directly spread from cell-to-cell (without contact with the extracellular milieu) provides a morphological explanation, why cellular immunity plays a crucial role in the immune protection against listeriosis. The cellular processes of L. monocytogenes infection have also been well characterized at the morphological level. A key group of L. monocytogenes virulence genes and their specific functions in the intracellular infection process have been identified and characterized. In tissue culture models of infection, the following stages of infection can be defined 1) internalization of L. monocytogenes within the host cell; 2) bacterial escape from the host vacuole; 3) multiplication of the bacterium within the host cell cytoplasm and movement through the cytoplasm by virtue of bacterially-directed polymerization of host actin filaments; 4) bacterial movement to the host cell surface and extrusion of bacterial cells in pseudopod-like structures; 5) phagocytosis of these pseudopod-like structures by neighboring cells, followed by escape of the bacterium from the resulting double-membrane vacuole, thus allowing the cycle to repeat. Gene products essential for each step of the infection process have also been identified. Six L. monocytogenes virulence genes (prfA, plcA, hlyA, mpl, actA, and plcB) are located together in one virulence gene cluster. Additional L. monocytogenes virulence genes (e.g., inlA, inlB), which are not physically linked to this virulence gene cluster, have also been described.2
Generally, the most common clinical signs of listeriosis in ruminants include encephalitis, septicemia, and intrauterine infections, which may lead to abortion or to birth of weak and/or septicemic animals. Less common symptoms associated with L. monocytogenes infections are mastitis, iritis and keratoconjunctivitis. In addition to clinically affected animals, a significant number of animals may be asymptomatic carriers of L. monocytogenes, often shedding the organism in fecal material.4
L. monocytogenes is predominantly transmitted by the oral route. Invasion generally occurs in the intestinal tract with subsequent hematogenous spread, leading to septicemia and possibly transuterine infection. The pathogenesis of listerial encephalitis is still somewhat controversial. There are some indications that L. monocytogenes may enter through the oral mucous membranes (possibly through pre-formed lesions) and subsequently migrate centripetally inside cranial nerves to the brain and particularly to the brainstem. For example, oral lesions in animals changing teeth may provide an entry port for L. monocytogenes.
Clinical Signs in Sheep
Widespread listeriosis in sheep was first recognized in 1931 in New Zealand. Encephalitic listeriosis (often referred to as "circling disease") is a result of brain stem lesions caused by L. monocytogenes. Common signs of encephalitis include fever, depression, lack of feed and water intake, dullness, turning or twisting of the head to one side, and movement in circles. In advanced cases unilateral facial nerve paralysis can develop, causing drooping of the eyelid and ear, and drooling. Encephalitis has been reported as the predominant symptom of listeriosis seen in sheep; it has been observed in lambs as young as 4-8 months. The incubation period can be up to 3 weeks and in most cases of listerial encephalitis in sheep, death occurs within two to three days of onset of clinical signs.2
Abortion represents another common clinical manifestation of listeriosis in sheep. Clinical signs in the ewe are resolved following abortion of the fetus, which usually occurs as a stillbirth in the third trimester of pregnancy. Septicemia is the most frequent infection type in neonates and lambs. Often, when a dam has a subclinical listerial infection she will bear an apparently healthy lamb that will then become septicemic and die within a few days. This infection is thought to be a consequence of intrauterine infection and is often characterized by diarrhea. Other clinical signs associated with listerial septicemia include elevated body temperatures and a loss of appetite. Septicemia is also often accompanied by hepatitis, splenitis and pneumonitis. Although the response to antibiotics is good in adult septicemic cases, pregnant animals may still abort following clinical recovery. Overall, the mortality rate is much lower for septicemic than for encephalitic cases. It has been reported that up to 5-10% of sheep exposed to L. monocytogenes during an outbreak show clinical signs, and many more are likely to have subclinical infections, shedding the bacteria in their feces, consistent with recent data that show that a high prevalence of L. monocytogenes in fecal, soil, feed, and water samples collected in small ruminant herds with listeriosis.5
Clinical Signs in Goats
The clinical manifestations of listeriosis in goats closely resemble those seen in sheep; encephalitis, septicemia, and abortion. In cases of listerial encephalitis in goats, meningitis frequently develops secondary to encephalitic brain stem lesions. As in ovine cases, goats usually succumb to an encephalitic listerial infection within two to three days, with fatalities reaching sixty percent. Septicemia is also often seen in goats, initial clinical signs include general depression, loss of appetite, a decrease in milk yield, and an increase in body temperature. Affected goats may also have diarrhea. In pregnant does, L. monocytogenes may cause abortion. Like sheep, goats may be asymptomatic carriers, shedding L. monocytogenes in the feces and milk. This provides the potential for environmental contamination and infection of newborn kids housed with the does through the navel or through sucking on dirty teats.2
Clinical Signs in Cattle
Some sources report that cattle account for about eighty to ninety percent of reported listeriosis cases in North America. This may, though, not necessarily reflect a higher true incidence of listeriosis in cattle, but could also reflect a lower reporting rate for listeriosis cases in small ruminants. Encephalitis, abortion, septicemia, and mastitis due to L. monocytogenes have been documented in cattle.6 In an encephalitic infection, death usually occurs about two weeks after the appearance of the first clinical signs, in contrast to the very rapid death seen in sheep. Bovine encephalitic listeriosis is characterized by walking in circles, bending the head to one side, with the corresponding ear drooping, the tongue protruding from the mouth, and abundant nasal secretion and drooling. Finally, the animal is increasingly emaciated, irritable, and then comatose until death. Neural damage to cranial nerves may also cause general irritability of the affected animal and impaired locomotion. Some animals that survive the listerial CNS infections show typical postencephalitic or postmyelitic symptoms like "dumbness", difficulty locating feed, and occasionally falling down, all results of the neural damage caused by the bacteria. The clinical signs of encephalitic listeriosis are similar to those associated with rabies or lead poisoning in cattle. Thus, it is important to consider listeriosis as a differential diagnosis in suspect cases of rabies or lead poisoning.
Bovine listerial septicemia is characterized by the same clinical signs seen in sheep and goats; elevated body temperature, a loss of appetite, and diarrhea. In cattle and particularly in newborn calves, septicemia is often accompanied by miliary abscesses, especially in the liver. Like in sheep, intrauterine infection may lead to the birth of septicemic calves; typical clinical signs in these calves include diarrhea. Listeriosis in cattle is also often associated with abortion, generally occurring in the final trimester of pregnancy.
Less common manifestations of listeriosis in cattle include mastitis, keratoconjunctivitis, and iritis. Listerial mastitis is characterized by shedding of L. monocytogenes in the milk, often either accompanied by a general septicemia or without any other clinical signs. In mastitic cows, the condition may be chronic, with the animal shedding the bacteria in her milk intermittently for up to twelve months. An interesting note is that healthy calves can be born to chronic carriers who shed L. monocytogenes in their milk. Listerial keratoconjunctivitis and iritis are thought to be caused by direct contact between the eye and contaminated silage, e.g., when an animal reaches its head into the feed. These symptoms do not only occur in cattle, but also in small ruminants and other animals, including horses.7
Transmission of L. Monocytogenes in Farm Animals
In farm animals, listeriosis is typically linked to consumption of poor quality silage that is contaminated with high levels of L. monocytogenes. Improperly fermented silage or pockets of improper fermentation with a high pH (>6.0) allow for multiplication of Listeria and some poorly fermented silages have shown Listeria contamination in excess of 108 cfu/g wet weight of silage. Application of molecular subtyping methods has been used in a number of listeriosis outbreaks in ruminants to characterize sources and transmission of L. monocytogenes. In many cases matching L. monocytogenes subtypes have been found in silage and in diseased animals as well as in fecal samples from asymptomatic animals.5,8 Interestingly, while L. monocytogenes prevalence is similarly high in fecal, feed, water and soil samples collected on dairy farms with and without a history of listeriosis (24.4 and 20.2%, respectively), prevalence is considerable higher in samples collected from small ruminant farms with a history of listeriosis (32.9%) as compared to those without a history of listeriosis (5.9%).5 These data indicate common exposure of dairy cows to L. monocytogenes (with disease possibly reflecting increased susceptibility of animals), while exposure may be less common in small ruminants. While listeriosis cases have also been described in animals not exposed to silage (particularly in goats), the source of infection in these cases has usually not been identified.
Clinical diagnosis of listeriosis in dairy animals is generally difficult as a variety of inflammatory and infectious diseases can cause clinical signs similar to those of listerial encephalitis. Many diseases also cause clinical signs identical to those of listerial septicemia and abortion. Important differential diagnoses for listerial encephalitis include rabies, viral CNS infections, polioencephalomalacia, lead poisoning and other intoxications. In general, definitive diagnosis of listeriosis can be achieved by bacterial culture (i.e., isolation of the organism from generally sterile tissue or body fluid samples such as CSF or blood), by histopathology and sometimes by serology. In most cases definitive diagnosis of listeriosis cannot be accomplished in a live animal, but requires a post-mortem pathological and histopathological examination and/or microbial culture on organs collected post-mortem.2
Listeriosis is often a rapidly progressing disease; in encephalitic cases death usually occurs within 2 days to 2 weeks of the first onset of clinical signs. Therefore, rapid diagnosis and treatment initiation immediately after onset of clinical signs are necessary for successful treatment. Since early diagnosis prior to death is rarely achieved, appropriate treatment is often initiated late and unsuccessfully.
Listeria is a facultative intracellular pathogen, thus the organism may be protected from the action of some antibiotics. Surprisingly, some antibiotics that do not penetrate intracellularly, still seem to be effective for treatment of listeriosis. Ampicillin and amoxicillin are often recommended as effective antibiotics for the treatment of listeriosis. Oxytetracycline and penicillin G also have been used successfully. All of these antibiotics, however, require administration of very high doses to be effective. For example, the recommended oxytetracycline treatment scheme requires doses as high as 10mg/kg body weight per day for at least five days. Penicillin G should be given at 44,000U/kg body weight daily for 1-2 weeks in order to be effective. In addition to antibiotic therapy, supportive therapy may be necessary. This may include fluid and electrolyte replacement for animals having difficulty eating and drinking as a result of neurological damage. Acidosis caused by excessive salivation can be treated by intravenous replacement of bicarbonate ions.2
Control and Prevention
As therapy of listeriosis in ruminants is often difficult and unsuccessful, prevention and prophylaxis play a critical role in the control of this disease. In most listeriosis outbreaks, particularly in cattle and sheep, silage appears to be the most likely source of infection. L. monocytogenes is a ubiquitous organism and is likely to naturally occur in plant materials used for silage preparation and/or in contaminating soil or other materials. Fecal contamination from birds and other animals can also contribute to Listeria contamination of silage. While initial contamination of ensiled materials is probably common, multiplication and survival of L. monocytogenes depends on the quality of silage fermentation. Since listerial growth is inhibited by a pH lower than about 5.0-5.5, Listeria does not multiply in properly prepared silage. Careful preparation and use of silage is thus critical for controlling listeriosis in ruminants. Emphasis should be placed on reducing the likelihood of multiplication of Listeria within a bale or bunk of silage, including through effective compaction and sealing, which will help prevent air pockets where Listeria could thrive.2
When feeding silage, additional precautions can be taken to help prevent listeriosis. Mold growth on silage indicates conditions that also allow growth of Listeria. Thus, any obviously moldy material should not be fed to animals and should be discarded. In particularly susceptible animal populations and in flocks or herds with recent listeriosis cases extra care should to be taken to feed only high quality silage and to discard improperly fermented silage. For example, ripped bales represent a particular risk and silage close to the top, sides, and front in a bunker silo may often be improperly fermented. In cases of listerial keratoconjunctivitis and iritis, measures to avoid direct eye contact with contaminated silage may minimize additional cases. The majority of listeriosis cases in dairy animals appear to occur from November to May, with a peak around February and March. This temporal distribution may be related to common feeding of silage rather than pasture during these times, with an increase in the likelihood of feeding poor quality and older silage later in the season. Thus, prophylactic and preventive measures are particularly important during certain times of the year.
Natural immunity to Listeria likely provides protection for most animals exposed to L. monocytogenes. Vaccination of susceptible populations appears to provide a means of minimizing the risk of significant health problems and subsequent economic losses caused by listeriosis outbreaks. For the present time, however, vaccination is not a valid option in the United States or the U.K., because there is no licensed vaccine available. A live, attenuated vaccine is available in some European countries and is claimed to be effective for sheep and goats. As L. monocytogenes is an intracellular pathogen, only live vaccines are likely to induce protective cell-mediated immunity. Killed vaccines are unlikely to lead to a protective immune response.2
1. Farber JM, Peterkin PI. Microbiol Rev 1991; 55:476.
2. Wiedmann M, Evans KE. pp. 777-782. In: Encyclopedia of Dairy Science, 2002, Academic Press, London.
3. Edelson BT, Unanue ER. Curr Opin Immun 2000; 12: 425.
4. Wesley IV. In: Ryser ET and Marth EH (eds.) Listeria, Listeriosis and Food Safety. pp. 39-73. 1999. Marcel Dekker, Inc. New York.
5. Nightingale KK et al. Appl. Environ. Microbiol 2004; 70 4458.
6. Rebhun WC, deLahunta A. JAVMA 1982; 180:395.
7. Evans K, et al. J Vet Diagn Invest 2004; 16:464.
8. Wiedmann M, et al. J Clin Microbiol 1996; 34:1086.