REVIEWS DEVELOPING ANIMAL MODELS FOR POLYMICROBIAL DISEASES Lauren O. Bakaletz Polymicrobial diseases involve two or more microorganisms that act synergistically, or in succession, to mediate complex disease processes. Although polymicrobial diseases in animals and humans can be caused by similar organisms, these disease are often also caused by organisms from different kingdoms, genera, species, strains, substrains and even by phenotypic variants of a single species. Animal models are often required to understand the mechanisms of pathogenesis, and to develop therapies and prevention regimes. However, reproducing polymicrobial diseases of humans in animal hosts presents significant challenges. There is now compelling evidence that many infectious also have important advantages over in vitro methods, diseases of humans (FIG. 1) and animals (TABLE 1) are including the presence of organized organ systems, an caused by more than one microorganism. The mixed intact immune system and, in inbred mice, specific microbial nature of these diseases has been recognized genetic backgrounds, and the availability of many since the early 1920s but there has been renewed interest reagents for characterizing the immune response to in this topic since the 1980s1,signalled by the publication sequential or co-infecting microorganisms. The availabil- of four important reviews (REFS 2–5) from 1982 to the pre- ity of mice with specific genetic backgrounds can have a sent date. Polymicrobial diseases (see BOX 1 for nomen- pivotal role in understanding the mechanisms of patho- clature) can be caused by the synergistic or sequential genesis of polymicrobial diseases, as exemplified by stud- action of infectious agents from either the same or dif- ies on septic peritonitis11–17,periodontal disease18 and ferent kingdoms, genera, species, strains or substrains, or Lyme arthritis19,20.Understanding the molecular mecha- by different phenotypic variants of a single species6. nisms underlying polymicrobial diseases of veterinary Polymicrobial diseases share underlying mechanisms of importance has also been facilitated by the use of animal pathogenesis, such as common predisposing factors models. These veterinary systems are useful examples for (BOX 2),but each disease has unique aspects. Although those researchers attempting to develop animal models the molecular mechanisms of some polymicrobial infec- of complex human diseases. tions are known, other polymicrobial diseases are not So far, most animal models for human polymicro- well understood. Owing to their complexity, the study of bial diseases are rodents, usually mice, but also rats, Center for Microbial polymicrobial infections requires a multidisciplinary gerbils, cotton rats and chinchillas. Other animal Pathogenesis, Columbus approach and specific in vitro methodologies and ani- models include non-human primates, which are use- Children’s Research mal models. The development of assay systems and ful for modelling diseases that are caused by microor- Institute, Department of treatment and prevention regimes is needed. ganisms with a restricted host range. For most human Pediatrics, The Ohio State University College of Multiple diverse in vitro systems have been used to viral co-infections of clinical importance, good ani- Medicine & Public Health, study polymicrobial diseases (BOX 3).Although in vitro mal models and culture systems are lacking and are 700 Children’s Drive, methods are crucial for understanding polymicrobial urgently required. Columbus, Ohio 43205- diseases, rigorous, reproducible and relevant animal This review provides an overview of the pathogenesis 2696, USA. models of human diseases are essential for the prevention of selected polymicrobial diseases, the molecular basis for e-mail: BakaletL@ 7–10 pediatrics.ohio-state.edu and treatment of these co-infections .All animal mod- some of these co-infections and describes animal models doi:10.1038/nrmicro928 els of human diseases have inherent limitations but they that have been developed to mimic these diseases. 552 | JULY 2004 | VOLUME 2 www.nature.com/reviews/micro REVIEWS Central nervous Table 1 | Examples of polymicrobial diseases system Sinuses • Multiple sclerosis* Host Infection or disease • Sinusitis • Alzheimer's disease* Cattle Bovine respiratory disease complex (BRDC), bovine gastroenteritis Ear Oral cavity Sheep Ovine conjunctivitis, ovine foot rot, chronic • Dental caries • Otitis media non-progressive pneumonia • Periodontitis • Adenotonsillitis Pigs Porcine atrophic rhinitis, porcine Respiratory tract gastroenteritis, porcine respiratory disease • Otitis media complex (PRDC) Heart • Pneumonia Poultry Poult enteritis mortality syndrome, • Stenotic artery • Pertussis disease infectious coryza • Atherosclerosis* • Kawasaki disease* Lung • Carotid coronary* • Cystic fibrosis • Pertussis the BVDV biotypes and can result in severe respiratory, • Chronic obstructive enteric or reproductive disease. The severity of the dis- pulmonary disease Gastrointestinal tract ease depends on the relative virulence of the viral strain, • Peritontitis physical and environmental stresses, and co-infection • Gastroenteritis Liver with another pathogen. • Hepatitis • Sarcoidosis* Bladder Porcine co-viral diseases. Porcine reproductive and respi- ratory syndrome (PRRS) results from infection with the Genitourinary tract • UTIs PRRS virus (PRRSV) followed by infection with a bacte- • Vaginitis rial or viral co-pathogen. Co-infecting viruses include • Inflammatory bowel porcine coronavirus, swine influenza virus22–24 and disease* pseudorabies virus25.Different plaque variants of PRRSV can also co-infect a porcine host26.Porcine post-weaning Skin and skeletal systems multisystemic wasting syndrome (PMWS) is due to • Necrotizing fasciitis • Group A streptococcal co-infection of pigs with porcine circovirus 2 (PCV-2) infection and porcine parvovirus (PPV)27,28. PCV-2 and PPV are • Lyme disease • Abscess thought to enter the host through tonsillar macrophages • Osteomyelitis and viraemia results within 3 days of infection. PCV-2 • Sarcoidosis* and PPV can replicate in circulating peripheral mono- cytes and contribute to both cell-associated viraemia and viral distribution throughout lymphoid tissues28. Hepatitis virus co-infection. Human co-infections with multiple hepatotropic viruses from the hepatitis virus group are well documented. Co-infection with multiple hepatitis viruses is possible owing to their similar routes of transmission and ability to chronically infect the host. Figure 1 | Human polymicrobial diseases. Polymicrobial diseases and suspected polymicrobial Hepatitis A virus (HAV) co-infection of individuals that diseases (indicated by an asterisk) are listed in the anatomical niche in which the disease are chronically infected with hepatitis B virus (HBV) pathology is mainly observed. and/or hepatitis C virus (HCV) results in a disease of increased severity and risk of death. Moreover, Viral co-infections HBV–HCV co-infection occurs in 10–15% of HBV Infections involving bovine viral diarrhoea virus. In cat- patients, and hepatitis G virus (HGV)–HCV co-infec- tle, infections with bovine viral diarrhoea virus (BVDV) tion occurs in 10–20% of individuals with chronic HCV can be clinically asymptomatic or can cause severe infection; however, HBV–hepatitis D virus (HDV) co- symptoms. The outcome depends on whether the pri- infection occurs only in the setting of co-infecting HBV. mary infection occurred in utero or after birth21 and VIRAL INTERFERENCE, in which replication of one virus is whether the primary infection was with a cytopathic or suppressed by another virus, is an intriguing aspect of non-cytopathic biotype of BVDV.Milder, congenital triple HBV–HCV–HDV infection — HDV can suppress persistent infection follows foetal infection with non- both HBV and HCV replication29.In a retrospective cytopathic BVDV. Death, or culling from the herd study of patients with hepatitis virus co-infections, HDV within 1 year of birth after failure to thrive, is common; was dominant by RT-PCR detection of HDV RNA in however some persistently infected calves seem healthy triple co-infections, but in dual co-infections there were at birth and survive for several years. Mucosal disease alternating dominant roles for either HBV or HCV. follows congenital persistent infection and is due to co- Multiple hepatotropic viral infections are associated with infection with cytopathic and non-cytopathic BVDV in reduced HCV replication but increased pathology. VIRAL INTERFERENCE One virus suppresses the utero. Conversely, acute bovine diarrhoeal disease is Patients with dual or triple co-infections have more replication of another. induced by primary post-natal infection with either of severe liver disease pathologies than patients that are NATURE REVIEWS | MICROBIOLOGY VOLUME 2 | JULY 2004 | 553 REVIEWS Co-infection with HIV and herpes simplex virus. In Box 1 | Nomenclature of polymicrobial diseases 1994, Kaposi’s sarcoma-associated herpesvirus For diseases and infections that involve two or more microorganisms, several terms are used (KSAV), also known as human herpesvirus type 8 in the literature, often interchangeably, and include: polymicrobial diseases, complex (HHSV-8), was isolated. KSAV is found in all clinical diseases, complicated infections, co-infections, concurrent infection, polybacterial diseases, variants of Kaposi’s sarcoma and is also associated dual infections, mixed infections, synergistic infections, superinfections and secondary with