The gram-positive bacterium Listeria monocytogenes is the causative agent of listeriosis, a highly fatal opportunistic foodborne infection. Pregnant women, neonates, the elderly, and debilitated or immunocompromised patients in general are predominantly affected, although the disease can also develop in normal individuals. Clinical manifestations of invasive listeriosis are usually severe and include abortion, sepsis, and meningoencephalitis. Listeriosis can also manifest as a febrile gastroenteritis syndrome. In addition to humans, L. monocytogenes affects many vertebrate species, including birds. Listeria ivanovii, a second pathogenic species of the genus, is specific for ruminants. Our current view of the pathophysiology of listeriosis derives largely from studies with the mouse infection model. Pathogenic listeriae enter the host primarily through the intestine. The liver is thought to be their first target organ after intestinal translocation. In the liver, listeriae actively multiply until the infection is controlled by a cell-mediated immune response. This initial, subclinical step of listeriosis is thought to be common due to the frequent presence of pathogenic L. monocytogenes in food. In normal indivuals, the continual exposure to listerial antigens probably contributes to the maintenance of anti-Listeria memory T cells. However, in debilitated and immunocompromised patients, the unrestricted proliferation of listeriae in the liver may result in prolonged low-level bacteremia, leading to invasion of the preferred secondary target organs (the brain and the gravid uterus) and to overt clinical disease. L. monocytogenes and L. ivanovii are facultative intracellular parasites able to survive in macrophages and to invade a variety of normally nonphagocytic cells, such as epithelial cells, hepatocytes, and endothelial cells. In all these cell types, pathogenic listeriae go through an intracellular life cycle involving early escape from the phagocytic vacuole, rapid intracytoplasmic multiplication, bacterially induced actin-based motility, and direct spread to neighboring cells, in which they reinitiate the cycle. In this way, listeriae disseminate in host tissues sheltered from the humoral arm of the immune system. Over the last 15 years, a number of virulence factors involved in key steps of this intracellular life cycle have been identified. This review describes in detail the molecular determinants of Listeria virulence and their mechanism of action and summarizes the current knowledge on the pathophysiology of listeriosis and the cell biology and host cell responses to Listeria infection. This article provides an updated perspective of the development of our understanding of Listeria pathogenesis from the first molecular genetic analyses of virulence mechanisms reported in 1985 until the start of the genomic era of Listeria research.

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Listeria pathogenesis and molecular virulence determinants.

Listeria monocytogenes virulence and pathogenicity, a food safety perspective.

https://www.ucipfg.com/Repositorio/MIA/MIA-07/Unidad_academica01/Control_of_L_monocytogenes.pdf

Control of Listeria monocytogenes in the Food-Processing Environment

The role of hemolysin in the virulence of Listeria monocytogenes was studied by using transposon mutagenesis. The 26-kilobase conjugative transposon Tn1545, originally found in Streptococcus pneumoniae, was transferred to a hemolytic virulent strain of L. monocytogenes. The frequency of transfer was estimated to be about 10(-8) per recipient. This allowed us to isolate a nonhemolytic mutant which most likely harbors a single copy of Tn1545. Loss of hemolysin production was associated with loss of virulence. The 50% lethal dose of the mutant was assessed to about 10(9.6) bacteria per mouse after intravenous challenge. Nonhemolytic bacteria were unable to grow in host tissues and were rapidly eliminated from the spleen and liver of infected mice. Virulence was restored in hemolysin-producing revertant obtained by spontaneous loss of transposon Tn1545. These results strongly suggest that hemolysin is a major virulence factor implicated in the intracellular growth of L. monocytogenes.

Transposon mutagenesis as a tool to study the role of hemolysin in the virulence of Listeria monocytogenes.

The role of listeriolysin O in the intracellular multiplication of Listeria monocytogenes and, therefore, its pathogenicity was questioned through a genetic complementation study. A nonhemolytic mutant was generated by inserting a single copy of transposon Tn917 in the bacterial chromosome. This insertion was localized by DNA sequence analysis in hlyA, the gene coding for listeriolysin O. As was another mutant that we previously characterized, this mutant was avirulent in the mouse. It was transformed with a plasmid carrying only hlyA, able to replicate in L. monocytogenes, and stably maintained in vitro and in vivo. The complemented strain displayed a hemolytic phenotype identical to that of the wild-type strain and was fully virulent, therefore attributing a crucial role to listeriolysin O in virulence and excluding the hypothesis of a polar effect of the transposon insertion on genes adjacent to hlyA and possibly involved in virulence.

https://iai.asm.org/content/iai/57/11/3629.full.pdf

Listeriolysin O is essential for virulence of Listeria monocytogenes: direct evidence obtained by gene complementation.

Different strains ofListeria monocytogenes serovar 1/2a were used to infect mice. A smooth, haemolytic strain multiplied in the spleen of normal adult mice and induced a long-lasting immunity to subsequent challenge infection. When the animals were treated with dextran sulphate (DS)Listeriae multiplied more rapidly and death followed within a few days. When normal baby mice were infected with this organism, fatal infection occurred. In nude mice a chronic infection developed. Secondly, a rough, haemolytic strain was used to infect mice. In normal adult mice no multiplication of this strain was observed. This holds true also for DS-treated animals. In nude mice the bacteria were eliminated slowly. Normal baby mice could only be killed if the infective dose was increased. This strain was considered to be intermediate in virulence. The infection stimulated a considerable immune response in mice, although to a lesser degree than the smooth, haemolyticListeria strain. Thirdly, a smooth, non-haemolytic strain ofL. monocytogenes serovar 1/2a was used. The bacteria were rapidly eliminated in normal, in DS-treated and in nude animals. Normal baby mice did not develop fatal disease, so it was considered that this strain ofL. monocytogenes serovar 1/2a was avirulent. This variant was found to be non-immunogenic. A boosting of immunity of animals by this avirulentListeria strain was, however, found to be possible. A reasonable explanation for the rapid elimination of avirulentL. monocytogenes serovar 1/2 a from mice cannot be presented at this time. Whereas it is known that the macrophage system and the T-lymphocytes play an essential role in the resistance to virulentListeriae, there is no increased susceptibility of the avirulent bacteria to these defence mechanisms. Other bacterial properties, such as serum sensitivity and lysozyme susceptibility, are likewise considered to be unimportant.

Virulence of different strains of Listeria monocytogenes serovar 1/2a.

Mice infected subcutaneously with 5 × 103 viable cells of Escherichia coli ATCC 25922 incorporated in liquified agar developed a systemic infection. Increasing bacterial numbers could be recovered from the liver at several days following infection. Ultimately, the animals died 6 days after infection.

The influence of killed Propionibacterium granulosum on experimental infection with Escherichia coli.

Mice immunized by primary infection with an avirulent strain ofToxoplasma gondii were protected against challenge infection with a highly virulent strain, even though macrophages were eliminated either by dextran sulfate or by carbon ink. This findings differs strikingly from previous results obtained in similar experiments with other intracellular pathogens such asListeria monocytogenes andSalmonella typhimurium. It is therefore concluded that the macrophage population, which plays an essential role in the cell-mediated immunity to reinfection withL.monocytogenes andS. typhimurium, is not of primary importance in cell-mediated immunity to reinfection withT.gondii.

The role of macrophages in acquired cell-mediated immunity to Toxoplasma gondii.

[Influence of Bordetella pertussis on the lymphatic tissue of mice. 8. The influence of Bordetella pertussis on the primary and secondary immune potential of thymusless ("nude") mice (author's transl)].

Verglichen mit phanotypisch normalen Kontrolltieren fanden sich bei NMRI-Mausen mit kongenitaler Thymusaplasie nur wenige Peyersche Plaques, die zudem nur aus diffusen Lymphocytenansammlungen bestanden. Auch die Milzfollikel waren erheblich verkleinert, wobei die periarteriolaren Follikelbereiche gelegentlich zellarmer erschienen. Das wei\e Blutbild der thymuslosen Tiere war durch eine ma\ige Lymphopenie charakterisiert. Obwohl die Milzen der Mause mit kongenitaler Thymusaplasie keine Verringerung der Zahlen an praexistenten 19S-Hamolysin-bildenden Zellen aufweisen, waren nach Erstimmunisierung mit 4·108 Schaferythrocyten nur minimale Zahlen an 19S-Produzenten und 7 S-Produzenten nachweisbar. Die sekundare antigene Stimulierung mit 4·108 Schaferythrocyten fuhrte zu keiner nennenswerten Gedachtnisreaktion, was anzeigt, da\ die Bildung von Gedachtniszellen ein thymusabhangiger Proze\ ist. Die alleinige intraperitoneale Injektion abgetoteter Zellen von B. pertussis fuhrte zu Hypersplenie, Leukozytose und Vermehrung der praexistenten Zahlen an 19S-Hamolysin-bildenden Zellen. Im Gegensatz zu Normaltieren, wo die Leukocytose durch eine Vermehrung von Granulocyten und Lymphocyten charakterisiert ist, fehlte bei den thymuslosen Tieren die Lymphocytenvermehrung. Der Zunahme der Blutgranulocyten entsprach eine verstarkte Granulopoese in Leber und Milz. Bei Normaltieren fand sich nach Erstimmunisierung durch simultane Applikation von 4·108 Schaferythrocyten und 3·109 abgetoteten Zellen vonB. pertussis eine verstarkte und verlangerte Produktion von 19S- und besonders 7S-Hamolysin-bildenden Zellen, verbunden mit einer verstarkten Praparation des lymphoretikularen Gewebes fur die anamnestische Reaktion. Bei den Mausen mit kongenitaler Thymusaplasie beschrankte sich der Adjuvanseffekt allem auf die Fruhphase der immunologischen Primarreaktion, was seinen Ausdruck in einer deutlichen Vermehrung der 19S-Produzenten und einer nur geringgradigen Vermehrung der 7S-Produzenten fand.

H. Hof

Papers

Virulence of different strains of Listeria monocytogenes serovar 1/2a.

The influence of killed Propionibacterium granulosum on experimental infection with Escherichia coli.

The role of macrophages in acquired cell-mediated immunity to Toxoplasma gondii.

[Influence of Bordetella pertussis on the lymphatic tissue of mice. 8. The influence of Bordetella pertussis on the primary and secondary immune potential of thymusless ("nude") mice (author's transl)].

Einflu\ vonBordetella pertussis auf das lymphatische Gewebe von Mäusen