Gram-negative Bacterial Infections

Overview of Gram-negative Bacteria

First Described: Gram-negative bacteria were first described in Berlin by the Danish scientist Hans Christian Gram in 1884 when he used Gram stain to visualize Klebsiella pneumoniae in the lungs of people who died of pneumonia.¹

Causes: Examples include Escherichia coli, Salmonella, Enterobacter, Citrobacter, Klebsiella, Campylobacter, Pseudomonas, and Acinetobacter.

Geographic Distribution: Worldwide

Mode of Transmission: Direct contact or contact with contaminated fomites; the bacteria often invade opportunistically

Major Clinical Signs: Affected sites include UTIs, wound infections, pneumonia, pyothorax, peritonitis, pyometra, ocular surface infections, otitis externa, pyoderma, bacteremia, and endocarditis, but virtually any body system can be affected.

Differential Diagnoses: Infections with gram-positive bacteria such as Staphylococcus spp. and Streptococcus spp.; to a lesser extent infections with nonbacterial pathogens such as fungi.

Human Health Significance: Gram-negative bacteria shed by dogs and cats have the potential to colonize humans, especially those with impaired host defenses. They may also be responsible for infections of dog or cat bites.

Etiology, Epidemiology, and Clinical Features

Gram-negative bacteria are widespread causes of opportunistic infections in dogs and cats and primarily cause disease when host defenses are impaired. They are important causes of nosocomial disease. In contrast to gram-positive bacteria, gram-negative bacteria have a complex outer membrane that contains lipopolysaccharide (LPS), as well as structures known as porins that regulate transport of molecules in and out of the cell, which includes antimicrobial drugs. Between this outer membrane and the inner (cytoplasmic) membrane is a periplasmic space, which contains the structural polymer peptidoglycan (or murein) (Figure 36-1). The peptidoglycan layer of gram-negative bacteria is much thinner than that of gram-positive bacteria, approximately one layer, or 7 to 8 nm thick (compared with many layers, or 20 to 80 nm thick for gram-positive bacteria). The inner cytoplasmic membrane contains many different proteins that are lipoproteins anchored into the outer lipid bilayer leaflet, transmembrane proteins, or peripheral membrane proteins that lie adjacent to the bilayer leaflets.

FIGURE 36-1 Structure of the envelope of gram-positive (A) and gram-negative (B) bacteria. The thickness of the peptidoglycan layer is greater in gram-positive bacteria than in gram-negative bacteria.

LPS is a large molecule that varies in composition from one bacterial species and strain to another. It contributes to the structural integrity of gram-negative bacteria and is a potent virulence factor. It consists of the lipid A backbone, a core oligosaccharide, and the O antigen side chain. Lipid A is a phosphorylated disaccharide to which long, hydrophobic fatty acid chains are attached, which anchor the LPS into the outer membrane (Figures 36-1 and 36-2). The lipid A component is also known as endotoxin because it is the biologically active portion of the molecule, in that it stimulates a potent host inflammatory response (see Chapter 86). The core oligosaccharide connects lipid A to the O antigen side chain. Its composition differs between bacterial species. The O antigen is a repeating polysaccharide that projects from the surface of the bacterial cell and varies in length from 1 to 60 repeats. It is the antigenic portion of the molecule and is responsible for serogroup classification of gram-negative bacteria.

FIGURE 36-2 Structure of bacterial lipopolysaccharide.

Gram-negative cocci, such as Moraxella or Neisseria, are not widely recognized as significant pathogens of dogs and cats. However, they are commensals of the oral cavity of dogs and cats, can cause bite wound infections in humans,² and Neisseria canis was isolated in pure culture from a deep mandibular abscess from one dog.³ Gram-negative rods are classified into Enterobacteriaceae and non-Enterobacteriaceae species. Enterobacteriaceae that cause disease in dogs and cats include Escherichia coli, Proteus, Salmonella, Enterobacter, Citrobacter, Serratia, and Klebsiella. Non-Enterobacteriaceae species include the Pasteurellaceae (Pasteurella multocida), as well as Pseudomonas aeruginosa and Acinetobacter. Other gram-negative bacteria are coccobacilli or spiral-shaped organisms and include Bartonella, Bordetella, Campylobacter, Francisella, Helicobacter, and Brucella.

Like gram-positive cocci, some gram-negative bacteria have tremendous ability to form biofilms, or bacterial aggregates that are embedded in a matrix of exopolysaccharide (bacterial slime), which form on living or nonliving surfaces. These protect the bacteria, and bacteria in biofilms can resist the effect of antimicrobial drugs and disinfectants.

Enterobacteriaceae

Many of the Enterobacteriaceae contribute to the normal gastrointestinal flora of animals. Organisms that belong to the Enterobacteriaceae possess a capsule (K antigen) and may be motile via flagella (H antigen). Some highly encapsulated species, such as Klebsiella and Enterobacter, have a mucoid appearance when they grow in culture. The flagella project from the bacterial cell and contain the filament-forming protein flagellin, which (along with LPS) can stimulate the host inflammatory response. The Enterobacteriaceae also possess fimbriae or pili, which are thinner than flagella and play a role in adhesion and sometimes bacterial aggregation. Finally, they possess a variety of complex secretion systems (type I, II, III secretion systems). These facilitate export of toxins such as hemolysins, and molecules that are inserted into, and form pores in, host cell membranes. The bacteria then use these pores to inject toxins directly into the host cell. The Enterobacteriaceae also have a variety of iron scavenging systems, which are important virulence factors, and transmissible plasmids that contain genes for virulence factors and antimicrobial resistance factors.

Escherichia coli

Escherichia coli is the most common cause of urinary tract infections (UTIs) in dogs and cats and can also be isolated from dogs and cats with pyometra, prostatitis, peritonitis, cholangitis, cholecystitis, bacteremia and endocarditis, bronchopneumonia (such as that secondary to aspiration), and pyothorax. There is one report of necrotizing fasciitis in a dog associated with E. coli infection.⁴

Antibiotic resistance in E. coli isolates from dogs and cats is an increasing problem; 51% of 376 isolates collected from sick dogs and cats in the United States in 2005 showed resistance to at least one drug, and 29% of the 376 isolates were multiple drug resistant (MDR; defined as resistant to three or more classes of antimicrobial drugs).⁵ The prevalence of resistance in 395 isolates from healthy dogs and cats from Canada was lower, 19%.⁶ Extended-spectrum β-lactamase enzyme production has also been identified in E. coli isolates from dogs and cats in the United States,⁷ and a few isolates were found to express New Dehli metallo-β-lactamase (NDM-1), which hydrolyses carbapenems.⁸ Nosocomial infections with MDR E. coli have also been described in dogs and cats.

In humans, strains of E. coli that cause UTIs differ from those that cause diarrhea, and there is some evidence that this may also be true for strains that cause UTIs versus pyometra in dogs and cats.9–11 Strains of E. coli that cause UTIs belong to the pathotype extraintestinal pathogenic E. coli (ExPEC). These strains possess a number of virulence factors, such as siderophores, type 1 and P fimbriae, hemolysin, and cytotoxic necrotizing factor. Specific virulence factors, as well as biofilm-forming capacity, have been associated with persistent or relapsing UTIs,^12,13 and uropathogenic E. coli can invade and persist within bladder epithelial cells. In one study, canine UTIs associated with E. coli were more likely to be recurrent than those associated with other uropathogens.¹⁴ There has been concern that dogs may act as a source of E. coli strains that are pathogenic for humans.^11,15 In other extraintestinal E. coli infections, E. coli more closely resemble commensal strains, and host factors may be more important than bacterial virulence factors. The reader is referred to Chapter 46 for more information on intestinal E. coli infections.

In dogs, S. marcescens infection has been associated with UTIs, bacteremia and aortic valve endocarditis,¹⁹ catheter-related infections in canine parvoviral enteritis,¹⁴ and necrotizing fasciitis.²⁰ Contamination of blood products was linked to an outbreak of bacteremia in cats.²¹ Serratia is resistant to ampicillin and first-generation cephalosporins as a result of a chromosomal β-lactamase. It may also acquire plasmid-mediated resistance to other antimicrobial drugs.