Where is listeriosis most commonly found?

Epidemiology

Listeria monocytogenes is widespread in nature, has been isolated throughout the environment, and is associated with epizootic disease and asymptomatic carriage in >42 species of wild and domestic animals and 22 avian species. Epizootic disease in large animals (e.g., sheep, cattle) is associated with abortion and circling disease, a form of basilar meningitis.L. monocytogenes is isolated from sewage, silage, and soil, where it survives for >295 days. Human-to-human transmission rarely occurs except in maternal-fetal transmission. The annual incidence of listeriosis decreased by 36% between 1996 and 2004 and has remained level since then. However, food-borne outbreaks continue to occur. In 2011, 84 cases and 15 deaths in 19 states were traced to cantaloupes from a single source. The cases were connected by use of pulsed-field gel electrophoresis, which showed that 4 different strains traced to the same source. The rate ofListeria infections varies among states. Epidemic human listeriosis has been associated with food-borne transmission in several large outbreaks, especially in association with aged soft cheeses; improperly pasteurized milk and milk products; contaminated raw and ready-to-eat beef, pork, and poultry, and packaged meats and salads; and vegetables both fresh and frozen harvested from farms where the ground is contaminated with the feces of colonized animals. Food-borne outbreaks in 2016 included raw milk, packaged salads, and frozen vegetables. The ability ofL. monocytogenes to grow at temperatures as low as 4°C (39.2°F) increases the risk for transmission from aged soft cheeses and stored contaminated food. Listeriosis is an uncommon but important recognized etiology of neonatal sepsis and meningitis. Small clusters of nosocomial person-to-person transmission have occurred in hospital nurseries and obstetric suites. Sporadic endemic listeriosis is less well characterized. Likely routes include food-borne infection and zoonotic spread.Zoonotic transmission with cutaneous infections occurs in veterinarians and farmers who handle sick animals.

Reported cases of listeriosis are clustered at the extremes of age. Some studies show higher rates in males and a seasonal predominance in the late summer and fall in the Northern hemisphere. Outside the newborn period and during pregnancy, disease is usually reported in patients with underlying immunosuppression, with a 100-300 times increased risk in HIV-infected persons and in the elderly population (Table 215.1). In a recent surveillance study from England, malignancies accounted for one third of cases, with special risk associated with cancer in elderly persons.

The incubation period, which is defined only for common-source food-borne disease, is 21-30 days but in some cases may be longer. Asymptomatic carriage and fecal excretion are reported in 15% of healthy persons and 5% of abattoir workers, but duration of excretion, when studied, is short (<1mo.).

Pathogenesis

L. monocytogenes is a hardy organism that is adapted to live as a saprophyte in external environments or as an intracellular pathogen in a number of animal hosts.30 Within its animal hosts, it maintains an arsenal of defensive mechanisms to survive in, respond to, and proliferate in the diverse environments it encounters.31L. monocytogenes can employ a number of virulence factors such as Internalin A (InlA) encoded byinlA and listeriolysin O (LLO) encoded byhly to help facilitate entry into and movement through the host cells, to escape phagosomes, and to circumvent the host immune response.32 Virulence genes are often used as targets for PCR detection and subtyping.33

In nearly all cases of infection,L. monocytogenes enters into human hosts via the gastrointestinal tract after ingestion of contaminated food. In the stomach, gastric pH of less than 3 is informally bactericidal; however bacterial survival increases at a pH of 3.5 and even more at a pH of 4.34 This is perhaps why the widespread use of proton pump inhibitors and thus elevated gastric pH has been shown to increase the risk of nonperinatal invasive listeriosis after adjusting for confounding factors.35L. monocytogenes then moves into the duodenum where the high concentration of bile creates a hostile environment for most microbes.36 The expression of bile salt hydrolases and the ability to tolerate high-salt conditions allowL. monocytogenes not only to survive transit through the duodenum but also to colonize the gallbladder.37,38 Luminal antibodies have not been shown to be protective againstL. monocytogenes, whereas innate immune mechanisms such as bactericidal peptides produced by Paneth and epithelial cells are successful host defenses in the small intestine.37

The intestinal microbiota also provides an independent and important first line of defense againstL. monocytogenes infection and indirectly augments host defenses.37,39 Potential mechanisms of protective properties of commensal bacteria are nutrient competition, contact-dependent inhibition and production of bacterial soluble mediators, and bacteriocins, which are toxic molecules created by certain common gut bacteria.37 Conversely, dysbiosis, from exposure to antibacterial agents or other causes, can allow for the expansion ofL. monocytogenes within the intestine. Listeriolysin S is a virulence factor found in epidemic strains ofL. monocytogenes and the only known bacteriocin produced in theListeria genus that disrupts the host gut microbiota creating a beneficial environment forL. monocytogenes during infection.40

Listeria monocytogenes first emerged as a serious threat to the dairy industry in 1985 when a major outbreak of listeriosis was traced to consumption of soft Mexican-style cheese in southern California, with other dairy-related outbreaks primarily involving pasteurized milk and certain soft cheeses prepared from raw milk having since been reported. Listeria monocytogenes causes abortion and perinatal septicemia in pregnant women and meningitis in the elderly and immunocompromised patients; it can also produce clinical and subclinical mastitis in ruminant animals, with about 2.55% of the raw milk supply found positive for this pathogen. Listeria monocytogenes is endemic to dairy farms and to a lesser extent dairy processing facilities where this pathogen is typically considered a postpasteurization contaminant. Unlike many other bacterial foodborne pathogens, L. monocytogenes can grow in milk at refrigeration temperatures and reach potentially infectious levels in certain high-moisture and surface-ripened cheeses. However, barring postpasteurization contamination, properly pasteurized fluid milk products will be free of Listeria.

Listeria monocytogenes

L. monocytogenes causes 2% to 8% of cases of bacterial meningitis in the United States and carries a mortality rate of 15% to 29%.4851 Serotypes 1/2b and 4b have been implicated in up to 80% of meningitis cases caused by this organism. Epidemiologic data show that the incidence ofL. monocytogenes meningitis is highest in neonates and elderly, with incidences up to 0.61 per 100,000 in neonates and 0.53 per 100,000 in the elderly.54 A study from the Netherlands showed that the incidence of neonatalListeria meningitis had decreased in the previous 25 years, potentially because of increased awareness of food restrictions for pregnant women. The rate of unfavorable outcome among adults withListeria meningitis was found to increase over a 14-year period from 27% to 61%, with the emergingL. monocytogenes sequence type 6 identified as the main factor leading to a poorer prognosis.115 A follow-up whole-genome sequencing study of theseListeria strains identified a plasmid containing the benzalkonium chloride tolerance gene that was associated with decreased susceptibility to disinfectants commonly used in the food-processing industry. Strains containing the plasmid had increased minimal inhibitory concentrations (MICs) to amoxicillin and gentamicin, which are commonly used in treatment ofL. monocytogenes infections.116

Listeria has been isolated from dust, soil, water, sewage, and decaying vegetable matter (including animal feed and silage). Listerial infection is most common in infants younger than 1 month (up to 10% of cases), adults older than 60 years, alcoholics, cancer patients, those receiving corticosteroid therapy, and immunosuppressed adults (e.g., renal transplant recipients).117119 Other predisposing conditions include diabetes mellitus, liver disease, chronic renal disease, collagen vascular diseases, pregnancy, and conditions associated with iron overload. Although colonization rates are low, pregnant women (who account for 25% of all cases of listeriosis) may harbor the organism asymptomatically in their genital tract and rectum and transmit the infection to their infants. Adults younger than 50 years withListeria meningitis should be screened for HIV infection.120 Meningitis can also occur in immunocompetent children and adults.121,122 Outbreaks ofListeria infection have been associated with the consumption of contaminated coleslaw, raw vegetables, milk, and cheese, with sporadic cases traced to contaminated turkey franks, alfalfa tablets, cantaloupe, diced celery, hog head cheese (a meat jelly made from hog heads and feet), and processed meats, thus pointing to the intestinal tract as the usual portal of entry.117119,123126

Characteristics of the Species

Although attention to this organism as a foodborne pathogen began in the 1980s, it initially was discovered almost 100 years ago. The original identification revealed its ability to survive intracellularly in monocytes and neutrophils, and hence its original name Bacterium monocytogenes. After being named Listerella hepatolytica in 1927, in 1940, the name was changed by J. H. H. Pirie again to L. monocytogenes to honor the surgeon Joseph Lister, who was also the namesake for the mouthwash Listerine.

Listeria monocytogenes is a Gram-positive non-spore-forming rod on the order of 0.52 μm in length. The Gram stain result becomes variable as the culture ages. In direct smears that are Gram stained, the organism may appear to be almost coccoid-like, causing confusion with streptococci. Flagella may be produced between 20 and 25 °C but not at 37 °C. They are catalase positive and oxidase negative. It produces a β-hemolysin on blood agar plates, which is part of the CAMP (so named for Christie, Atkins, and Munch-Petersen) diagnostic test.

The organism can grow at temperatures ranging from <1 °C to approximately 50 °C, with an optimum temperature of 3037 °C. Listeria monocytogenes is quite hardy, and hence it is found frequently in the environment. The organism can withstand freezing, but it is inactivated by heating at 60 °C for 30 min. Heat inactivation was at one point a cause of concern as early claims were made that L. monocytogenes could survive pasteurization perhaps as a function of its ability to invade neutrophils and leukocytes in milk. Those reports subsequently were disavowed, and it became clear that L. monocytogenes could not survive standard pasteurization. It is a facultative anaerobe that can grow over a pH range of 49.5 and has the ability to grow in 10% sodium chloride. Acetate is the most effective acidulant in terms of inhibiting its growth. The minimum water activity for growth ranges from 0.90 to 0.97.

Listeria monocytogenes can ferment a number of carbohydrates including hexoses and pentoses with differences occurring depending upon aerobic versus anaerobic conditions. The carbohydrate fermentation patterns can be used to distinguish it from other Listeria spp.

1 Introduction

Listeria monocytogenes is a remarkable bacterial pathogen not only because of the sophisticated molecular mechanisms that it uses to invade and colonize the mammalian host (Cossart, 2011; de las Heras, Cain, Bielecka, & Vazquez-Boland, 2011; Radoshevich & Cossart, 2018) but also because it is exquisitely well adapted to cope with a range of environmental challenges including osmotic and acid stresses as well as cold temperatures (Gandhi & Chikindas, 2007; NicAogain & O'Byrne, 2016; O'Byrne & Karatzas, 2008; van Schaik & Abee, 2005). The latter properties make this food-borne pathogen particularly difficult to eliminate from the food chain, especially in so-called ready-to-eat foods, those foods that can be eaten without prior cooking (NicAogain & O'Byrne, 2016). Although infections are not very common in healthy individuals, the high mortality rate associated with infections (de Noordhout et al., 2014; Lecuit, 2007) combined with the ubiquity of this organism in the environment mean that is taken very seriously by food producers, and it continues to represent a serious public health risk. A key step in developing improved food safety measures is to develop a mechanistic understanding of how this organism protects itself in the complex and challenging environments it encounters, both within the food chain and within the host. Such an understanding could then be used to inform the rational design of new control measures that target the Achilles heel of this pathogen, in order to prevent its survival and growth at critical points along the food chain.

A key step in adapting to new stresses in the environment is the reprogramming of the transcriptional landscape to align gene expression with the physiological needs of the cell, and this is achieved by a panoply of both protein and ribonucleic acid transcriptional regulators. At the top of the hierarchy of transcriptional regulation lie the sigma factors, which largely determine the genes that are transcribed at any time by directing the transcriptional machinery to the appropriate promoter sequences. Most L. monocytogenes strains have five sigma factors, including the principal housekeeping sigma factor σA and four alternative sigma factors, σB, σC, σH, and σL (Glaser et al., 2001; O'Byrne & Karatzas, 2008). σB is the factor that controls the general stress response in L. monocytogenes and of the four alternative sigma factors, it has the largest regulon with almost 300 genes (approximately 10% of the genome) under the positive control of this sigma factor (Chaturongakul et al., 2011). Wiedmann and colleagues identified the sigB locus based on homology with the σB in Bacillus subtilis and demonstrated an important role for this sigma factor in acid tolerance (Wiedmann, Arvik, Hurley, & Boor, 1998). Almost simultaneously Becker, Sevket, Etin, Hutkins, and Benson (1998) identified the same locus and showed the involvement of σB in the response to osmotic stress.

Subsequently, σB in L. monocytogenes has received a lot of research attention with several studies helping to define fully the regulon (Abram, Starr, et al., 2008, Abram, Su, et al., 2008; Kazmierczak, Mithoe, Boor, & Wiedmann, 2003; Raengpradub, Wiedmann, & Boor, 2008; Toledo-Arana et al., 2009; Wemekamp-Kamphuis et al., 2004; Wurtzel et al., 2012). Genes under σB control are known to contribute to a variety of stress resistance mechanisms including osmoregulation (Cetin, Zhang, Hutkins, & Benson, 2004; Fraser, Sue, Wiedmann, Boor, & O'Byrne, 2003; Sue, Boor, & Wiedmann, 2003), acid tolerance (Cotter, Gahan, & Hill, 2001; Wemekamp-Kamphuis et al., 2004; Wiedmann et al., 1998), bile tolerance (Begley, Sleator, Gahan, & Hill, 2005; Zhang et al., 2011), cell wall acting antimicrobials (Begley, Hill, & Ross, 2006), and visible light (O'Donoghue et al., 2016; Ondrusch & Kreft, 2011; Tiensuu, Andersson, Rydén, & Johansson, 2013). Its role in surviving the gastrointestinal phase of the infectious cycle is also now well established (Dowd, Joyce, Hill, & Gahan, 2011; Garner, Njaa, Wiedmann, & Boor, 2006; Sleator, Watson, Hill, & Gahan, 2009; Toledo-Arana et al., 2009). There is also substantial evidence that σB plays an important role in virulence. First, the gene encoding the virulence master regulator PrfA is preceded by two overlapping promoters, one of which is recognized by σB (Kazmierczak, Wiedmann, & Boor, 2006; Rauch, Luo, Muller-Altrock, & Goebel, 2005). Second, the inlAB operon that encodes the cell invasion proteins internalin A and B are under σB control (Kim, Gaidenko, & Price, 2004; Kim, Marquis, & Boor, 2005). Overall the current view is that σB plays a vital role in the early gastrointestinal stages of the infection whereas the regulator PrfA dominates during systemic spread and the intracellular stages of the infectious cycle (de las Heras et al., 2011; O'Byrne & Karatzas, 2008).

While there is certainly still lots to learn about the systems under σB control and the roles they play in survival of this pathogen in the environment, the biggest outstanding questions relate to the mechanisms that control the activation of σB. Understanding the details of how σB becomes activated will be a critical step in developing strategies to undermine its protective functions and ultimately to prevent this pathogen from surviving in the human and animal food chains. This review attempts to review our current understanding of the σB system in L. monocytogenes including the sensory mechanisms that trigger its activation. We also discuss the regulatory overlap between this sigma factor and other important global regulators, including small regulatory RNAs, in this pathogen. The role that it plays in both the saprophytic and virulence phases of the life cycle of L. monocytogenes are also presented, along with some of the outstanding questions and challenges in this field.

Abstract

Infection with Listeria monocytogenes shows an early stage of lymphocyte apoptosis. This is an obligatory stage the extent of which depends on infective dose. Lymphocyte apoptosis occurs early and is rapidly superseded, yet it has a strong biological consequence. The immunological effect of lymphocyte apoptosis following infection is increased susceptibility to L. monocytogenes infection due, in part, to upregulation of IL-10 on macrophages and DC. Lymphocyte apoptosis is dependent on bacterial expression of the pore-forming toxin listeriolysin O (LLO). Also, purified LLO can lead to the induction of death pathways similar to infection, demonstrating that it is a killer agent generated by L. monocytogenes. Signaling through the type I interferon receptor potentiates cell death induced by the bacteria or LLO. Infection with L. monocytogenes also causes death of phagocytic cells, the nature and significance of which is not clear at present. Infection with L. monocytogenes is a tractable model to examine pathogen-induced cell death pathways and their possible immunological consequences in multiple cell types following infection.

Basic Information