Cross-species transmission of a novel adenovirus associated with a fulminant pneumonia outbreak in a new world monkey colony.

Adenoviruses are DNA viruses that naturally infect many vertebrates, including humans and monkeys, and cause a wide range of clinical illnesses in humans. Infection from individual strains has conventionally been thought to be species-specific. Here we applied the Virochip, a pan-viral microarray, to identify a novel adenovirus (TMAdV, titi monkey adenovirus) as the cause of a deadly outbreak in a closed colony of New World monkeys (titi monkeys; Callicebus cupreus) at the California National Primate Research Center (CNPRC). Among 65 titi monkeys housed in a building, 23 (34%) developed upper respiratory symptoms that progressed to fulminant pneumonia and hepatitis, and 19 of 23 monkeys, or 83% of those infected, died or were humanely euthanized. Whole-genome sequencing of TMAdV revealed that this adenovirus is a new species and highly divergent, sharing <57% pairwise nucleotide identity with other adenoviruses. Cultivation of TMAdV was successful in a human A549 lung adenocarcinoma cell line, but not in primary or established monkey kidney cells. At the onset of the outbreak, the researcher in closest contact with the monkeys developed an acute respiratory illness, with symptoms persisting for 4 weeks, and had a convalescent serum sample seropositive for TMAdV. A clinically ill family member, despite having no contact with the CNPRC, also tested positive, and screening of a set of 81 random adult blood donors from the Western United States detected TMAdV-specific neutralizing antibodies in 2 individuals (2/81, or 2.5%). These findings raise the possibility of zoonotic infection by TMAdV and human-to-human transmission of the virus in the population. Given the unusually high case fatality rate from the outbreak (83%), it is unlikely that titi monkeys are the native host species for TMAdV, and the natural reservoir of the virus is still unknown. The discovery of TMAdV, a novel adenovirus with the capacity to infect both monkeys and humans, suggests that adenoviruses should be monitored closely as potential causes of cross-species outbreaks.

Identification of cardioviruses related to Theiler's murine encephalomyelitis virus in human infections.

Cardioviruses comprise a genus of picornaviruses that cause severe illnesses in rodents, but little is known about the prevalence, diversity, or spectrum of disease of such agents among humans. A single cardiovirus isolate, Saffold virus, was cultured in 1981 in stool from an infant with fever. Here, we describe the identification of a group of human cardioviruses that have been cloned directly from patient specimens, the first of which was detected using a pan-viral microarray in respiratory secretions from a child with influenza-like illness. Phylogenetic analysis of the nearly complete viral genome (7961 bp) revealed that this virus belongs to the Theiler's murine encephalomyelitis virus (TMEV) subgroup of cardioviruses and is most closely related to Saffold virus. Subsequent screening by RT-PCR of 719 additional respiratory specimens [637 (89%) from patients with acute respiratory illness] and 400 cerebrospinal fluid specimens from patients with neurological disease (aseptic meningitis, encephalitis, and multiple sclerosis) revealed no evidence of cardiovirus infection. However, screening of 751 stool specimens from 498 individuals in a gastroenteritis cohort resulted in the detection of 6 additional cardioviruses (1.2%). Although all 8 human cardioviruses (including Saffold virus) clustered together by phylogenetic analysis, significant sequence diversity was observed in the VP1 gene (66.9%-100% pairwise amino acid identities). These findings suggest that there exists a diverse group of novel human Theiler's murine encephalomyelitis virus-like cardioviruses that hitherto have gone largely undetected, are found primarily in the gastrointestinal tract, can be shed asymptomatically, and have potential links to enteric and extraintestinal disease.

Rapid influenza antigen test for diagnosis of pandemic (H1N1) 2009.

We compared the QuickVue Influenza test with PCR for diagnosing pandemic (H1N1) 2009 in 404 persons with influenza-like illness. Overall sensitivity, specificity, and positive and negative predictive values were 66%, 84%, 84%, and 64%, respectively. Rapid test results should be interpreted cautiously when pandemic (H1N1) 2009 virus is suspected.

Rapid identification of known and new RNA viruses from animal tissues.

Viral surveillance programs or diagnostic labs occasionally obtain infectious samples that fail to be typed by available cell culture, serological, or nucleic acid tests. Five such samples, originating from insect pools, skunk brain, human feces and sewer effluent, collected between 1955 and 1980, resulted in pathology when inoculated into suckling mice. In this study, sequence-independent amplification of partially purified viral nucleic acids and small scale shotgun sequencing was used on mouse brain and muscle tissues. A single viral agent was identified in each sample. For each virus, between 16% to 57% of the viral genome was acquired by sequencing only 42-108 plasmid inserts. Viruses derived from human feces or sewer effluent belonged to the Picornaviridae family and showed between 80% to 91% amino acid identities to known picornaviruses. The complete polyprotein sequence of one virus showed strong similarity to a simian picornavirus sequence in the provisional Sapelovirus genus. Insects and skunk derived viral sequences exhibited amino acid identities ranging from 25% to 98% to the segmented genomes of viruses within the Reoviridae family. Two isolates were highly divergent: one is potentially a new species within the orthoreovirus genus, and the other is a new species within the orbivirus genus. We demonstrate that a simple, inexpensive, and rapid metagenomics approach is effective for identifying known and highly divergent new viruses in homogenized tissues of acutely infected mice.

Rhinovirus associated with severe lower respiratory tract infections in children.

Rhinovirus is a respiratory virus most typically associated with the common cold and asthma exacerbations, and has not traditionally been considered to play a major role in severe lower respiratory tract infections (LRTIs). As part of a surveillance program for respiratory pathogens of public health importance, children consecutively admitted to intensive care for LRTI at a large tertiary children's hospital were tested with polymerase chain reaction for 11 respiratory viruses and Mycoplasma pneumoniae from February 21 to October 31, 2007; 43 cases were enrolled and rhinovirus was the most frequently detected pathogen, with 21 (49%) positive. Rhinovirus cases frequently were young (median age, 1.4 years [range, 44 days-15 years]), hospitalized for pneumonia (10; 48%), had chronic underlying illnesses (15; 71%), had abnormal chest radiographs (18; 86%), required mechanical ventilation (12; 57%), and had prolonged hospitalization (median length, 7 days [range, 1-29 days]). Coinfection with other viruses or bacteria was common (10; 47%). Rhinovirus may be associated with more severe LRTI in children than previously reported, particularly in the noninfluenza, nonrespiratory syncytial virus season.

Severe pneumonia due to adenovirus serotype 14: a new respiratory threat?

BACKGROUND: Adenoviruses are associated with sporadic infection and community and institutional outbreaks; they can cause especially severe disease in infants, young children, immunocompromised persons, and transplant recipients. Fifty-two adenovirus serotypes have been recognized and classified within 7 subgroups or species (A-G), with limited data available on associated clinical syndromes and disease severity in more than one-half of the known serotypes. METHODS: We describe the clinical presentation and virologic characterization of 1 adult and 2 pediatric patients admitted to 2 separate hospitals during April-May 2006 with severe acute respiratory tract infection. All patients had underlying chronic pulmonary disease; none were severely immunocompromised. All 3 experienced serious chronic sequelae or died. RESULTS: Adenovirus was isolated from all 3 case patients. Adenovirus serotype 14, a subspecies B2 serotype not previously associated with severe clinical illness, was confirmed by neutralization assay and sequencing of the hexon gene. Restriction enzyme analysis with BamHI, BglII, HindIII, and SmaI showed all 3 viruses to be identical and to belong to a new genome type that we have designated "Ad14a." CONCLUSIONS: Our identification of severe respiratory illness due to a previously rarely reported adenovirus serotype may signify the emergence in the United States of a new genomic variant that has the potential to spread globally and cause epidemics. These case reports highlight the need for rapid diagnosis and improved surveillance, with serotyping and molecular characterization, to identify emerging variants of adenovirus, which may assist with targeted development of antiviral agents or type-specific vaccines.

Assay for 5' noncoding region analysis of all human rhinovirus prototype strains.

Increasing recognition of the association of rhinovirus with severe lower respiratory tract illnesses has clarified the need to understand the relationship between specific serotypes of rhinovirus and their clinical consequences. To accomplish this, a specific and sensitive assay to detect and serotype rhinovirus directly from clinical specimens is needed. Traditional methods of serotyping using culture and serum neutralization are time-consuming, limited to certain reference laboratories, and complicated by the existence of over 100 serotypes of human rhinoviruses (HRVs). Accordingly, we have developed a sequence-based assay that targets a 390-bp fragment accounting for approximately two-thirds of the 5' noncoding region (NCR). Our goal was to develop an assay permitting amplification of target sequences directly from clinical specimens and distinction among all 101 prototype strains of rhinoviruses. We determined the sequences of all 101 prototype strains of HRV in this region to enable differentiation of virus genotypes in both viral isolates and clinical specimens. We evaluated this assay in a total of 101 clinical viral isolates and 24 clinical specimens and compared our findings to genotyping results using a different region of the HRV genome (the VP4-VP2 region). Five specimens associated with severe respiratory disease in children did not correlate with any known serotype of rhinovirus and were found to belong to a novel genogroup of rhinovirus, genogroup C. Isolates were also found that corresponded to the genogroup A2 variant identified in New York and Australia and two other novel group A clusters (GAC1 and GAC2).

Genotype prevalence and risk factors for severe clinical adenovirus infection, United States 2004-2006.

BACKGROUND: Recently, epidemiological and clinical data have revealed important changes with regard to clinical adenovirus infection, including alterations in antigenic presentation, geographical distribution, and virulence of the virus. METHODS: In an effort to better understand the epidemiology of clinical adenovirus infection in the United States, we adopted a new molecular adenovirus typing technique to study clinical adenovirus isolates collected from 22 medical facilities over a 25-month period during 2004-2006. A hexon gene sequence typing method was used to characterize 2237 clinical adenovirus-positive specimens, comparing their sequences with those of the 51 currently recognized prototype human adenovirus strains. In a blinded comparison, this method performed well and was much faster than the classic serologic typing method. RESULTS: Among civilians, the most prevalent adenovirus types were types 3 (prevalence, 34.6%), 2 (24.3%), 1 (17.7%), and 5 (5.3%). Among military trainees, the most prevalent types were types 4 (prevalence, 92.8%), 3 (2.6%), and 21 (2.4%). CONCLUSIONS: For both populations, we observed a statistically significant increasing trend of adenovirus type 21 detection over time. Among adenovirus isolates recovered from specimens from civilians, 50% were associated with hospitalization, 19.6% with a chronic disease condition, 11% with a bone marrow or solid organ transplantation, 7.4% with intensive care unit stay, and 4.2% with a cancer diagnosis. Multivariable risk factor modeling for adenovirus disease severity found that age <7 years (odds ratio [OR], 3.2; 95% confidence interval [CI], 1.4-7.4), chronic disease (OR, 3.6; 95% CI, 2.6-5.1), recent transplantation (OR, 2.7; 95% CI, 1.3-5.2), and adenovirus type 5 (OR, 2.7; 95% CI, 1.5-4.7) or type 21 infection (OR, 7.6; 95% CI, 2.6-22.3) increased the risk of severe disease.

Molecular characterization of a variant rhinovirus from an outbreak associated with uncommonly high mortality.

BACKGROUND: Human rhinoviruses (HRVs) are the most frequent cause of acute upper respiratory tract infection, however, they are also known to replicate in the lower respiratory tract and associate with more severe respiratory illnesses. An outbreak of HRV occurred in a long-term facility in Santa Cruz, California with unusually high morbidity and mortality. OBJECTIVES: To identify viral characteristics associated with this unique outbreak, genetic relationships between these clinical isolates (SCRVs) and prototype strains of rhinovirus were investigated. STUDY DESIGN: Sequence homology and phylogenetic analyses of the SCRV VP4/VP2 region were performed in conjunction with all HRV prototypes. Due to the importance of the 5'noncoding region (NCR) and the structural genes to viral replication and host immune responses, respectively, we focused on a segment of the HRV genome which includes these regions. Molecular models of SCRV were also assessed. RESULTS: SCRV showed closest similarity to HRV82 with some divergence from the prototype. Amino acid differences were concentrated within predicted neutralization epitopes within VP2, VP3 and VP1. CONCLUSION: Sequence analyses and differences in cell culture growth characteristics suggest that this virus is a variant of HRV which has distinctive properties from its respective prototype strain.

Genome-wide diversity and selective pressure in the human rhinovirus.

BACKGROUND: The human rhinoviruses (HRV) are one of the most common and diverse respiratory pathogens of humans. Over 100 distinct HRV serotypes are known, yet only 6 genomes are available. Due to the paucity of HRV genome sequence, little is known about the genetic diversity within HRV or the forces driving this diversity. Previous comparative genome sequence analyses indicate that recombination drives diversification in multiple genera of the picornavirus family, yet it remains unclear if this holds for HRV. RESULTS: To resolve this and gain insight into the forces driving diversification in HRV, we generated a representative set of 34 fully sequenced HRVs. Analysis of these genomes shows consistent phylogenies across the genome, conserved non-coding elements, and only limited recombination. However, spikes of genetic diversity at both the nucleotide and amino acid level are detectable within every locus of the genome. Despite this, the HRV genome as a whole is under purifying selective pressure, with islands of diversifying pressure in the VP1, VP2, and VP3 structural genes and two non-structural genes, the 3C protease and 3D polymerase. Mapping diversifying residues in these factors onto available 3-dimensional structures revealed the diversifying capsid residues partition to the external surface of the viral particle in statistically significant proximity to antigenic sites. Diversifying pressure in the pleconaril binding site is confined to a single residue known to confer drug resistance (VP1 191). In contrast, diversifying pressure in the non-structural genes is less clear, mapping both nearby and beyond characterized functional domains of these factors. CONCLUSION: This work provides a foundation for understanding HRV genetic diversity and insight into the underlying biology driving evolution in HRV. It expands our knowledge of the genome sequence space that HRV reference serotypes occupy and how the pattern of genetic diversity across HRV genomes differs from other picornaviruses. It also reveals evidence of diversifying selective pressure in both structural genes known to interact with the host immune system and in domains of unassigned function in the non-structural 3C and 3D genes, raising the possibility that diversification of undiscovered functions in these essential factors may influence HRV fitness and evolution.

Creating a model program for influenza surveillance in California: results from the 2005-2006 influenza season.

BACKGROUND: Influenza surveillance is valuable for monitoring trends in influenza-related morbidity and mortality. Using the 2005-2006 influenza season as an example, this paper describes a comprehensive influenza surveillance program used by the California Department of Public Health (CDPH). METHODS: Data collected from patients evaluated for acute respiratory illness in a given week were reported and summarized the following week, including (1) electronic hospital pneumonia and influenza admission and antiviral usage records from Kaiser Permanente, (2) sentinel provider influenza-like illness (ILI) reports, (3) severe pediatric influenza case reports (e.g., children either hospitalized in intensive care or expired), (4) school clinic ILI evaluations, and (5) positive influenza test results from a network of academic, hospital, commercial, and public health laboratories and the state CDPH Viral and Rickettsial Disease Laboratory. RESULTS: Influenza activity in California in the 2005-2006 season was moderate in severity; all clinical and laboratory markers rose and fell consistently. Extensive laboratory characterization identified the predominant circulating virus strain as A/California/7/2004(H3N2), which was a component of the 2005-2006 influenza vaccine; 96% of samples tested showed adamantane resistance. CONCLUSIONS: By using multiple, complementary surveillance methods coupled with a strong laboratory component, the CDPH has developed a simple, flexible, stable, and widely accepted influenza surveillance system that can monitor trends in statewide influenza activity, ascertain the correlation between circulating strains with vaccine strains, and assist with detection of new strain variants. The methods described can serve as a model for influenza surveillance in other states.

Rhinovirus outbreak in a long term care facility for elderly persons associated with unusually high mortality.

During a 6-week period in 2003, 56 residents and 26 staff developed respiratory illness in a long-term facility; 12 residents died. Seven of 13 respiratory specimens were culture-positive for rhinovirus; 6 of the isolates were serotype 82. In elderly populations, severe illness may be associated with organisms typically considered to be "benign," such as rhinovirus.

Adenovirus, adeno-associated virus and Kawasaki disease.

Clinical similarities and shared seasonality suggested a relationship between adenovirus infection and Kawasaki disease. We performed adenovirus serology and quantitative polymerase chain reaction for both adenovirus and adeno-associated virus in patients with acute Kawasaki disease. No evidence was found to suggest a link between either virus and Kawasaki disease.

Detection of infectious human adenoviruses in tertiary-treated and ultraviolet-disinfected wastewater.

A pilot-scale study investigating the use of low-pressure, high-intensity UV radiation for disinfection of urban wastewater was conducted. The inactivation of coliform bacteria, wastewater-indigenous enteric viruses, seeded poliovirus, and seeded F-specific coliphage was studied. During the course of the pilot study, infectious human adenoviruses were isolated from 15 of 16 large-volume samples of UV-disinfected secondary- and tertiary-treated wastewater. Half of the tertiary-treated, UV-disinfected effluent samples from which the adenoviruses were isolated had total coliform concentrations that complied with California's Water Recycling Criteria. To determine the relative UV resistance of the adenovirus isolates, purified viruses were seeded into tertiary-treated waste-water and exposed to low-pressure, high-intensity, collimated UV radiation. A dose of approximately 170 mW-s/cm2 was required to achieve 99.99% inactivation. These findings suggest that UV doses effective at meeting certain wastewater regulations for total coliform bacteria may not provide suitable inactivation of the UV-resistant human adenoviruses.

Experimental cross-species infection of common marmosets by titi monkey adenovirus.

Adenoviruses are DNA viruses that infect a number of vertebrate hosts and are associated with both sporadic and epidemic disease in humans. We previously identified a novel adenovirus, titi monkey adenovirus (TMAdV), as the cause of a fulminant pneumonia outbreak in a colony of titi monkeys (Callicebus cupreus) at a national primate center in 2009. Serological evidence of infection by TMAdV was also found in a human researcher at the facility and household family member, raising concerns for potential cross-species transmission of the virus. Here we present experimental evidence of cross-species TMAdV infection in common marmosets (Callithrix jacchus). Nasal inoculation of a cell cultured-adapted TMAdV strain into three marmosets produced an acute, mild respiratory illness characterized by low-grade fever, reduced activity, anorexia, and sneezing. An increase in virus-specific neutralization antibody titers accompanied the development of clinical signs. Although serially collected nasal swabs were positive for TMAdV for at least 8 days, all 3 infected marmosets spontaneously recovered by day 12 post-inoculation, and persistence of the virus in tissues could not be established. Thus, the pathogenesis of experimental inoculation of TMAdV in common marmosets resembled the mild, self-limiting respiratory infection typically seen in immunocompetent human hosts rather than the rapidly progressive, fatal pneumonia observed in 19 of 23 titi monkeys during the prior 2009 outbreak. These findings further establish the potential for adenovirus cross-species transmission and provide the basis for development of a monkey model useful for assessing the zoonotic potential of adenoviruses.

An algorithm for the typing of enteroviruses and correlation to serotyping by viral neutralization.

BACKGROUND: Human enteroviruses (HEVs) are common pathogens which cause a broad spectrum of illnesses ranging from asymptomatic infection to acute myocarditis and aseptic meningitis. The neutralization assay for serotype determination is labor-intensive and time-consuming. There is a need for a methodology that is more rapid and widely accessible. OBJECTIVES: Our goals were to develop an algorithm to type enteroviruses which combines both serologic typing, based on indirect immunofluorescence assay (IFA) using type-specific monoclonal antibodies (mAbs) and genotyping, by DNA sequence analysis and to assess the correlation of both IFA and genotyping to traditional viral neutralization by type-specific antisera. STUDY DESIGN: Clinical specimens initially determined to be enterovirus positive by nucleic acid detection were grown in cell culture and typed using mAbs. Specimens that could not be typed by mAbs were subject to molecular analysis. Genotyping was performed by a combination of either a primary or semi-nested RT-PCR for a region within VP3/VP1 and followed by direct DNA sequencing of PCR products. Database homology comparisons and phylogenetic analysis were performed based on a defined region (303 nt) within the VP1 gene. RESULTS: We inoculated 134 enterovirus nucleic acid amplification-positive specimens into culture and 115 (86%) of these isolates were successfully typed by this algorithm. We have demonstrated a strong correlation between serotyping by viral neutralization to both IFA by type-specific mAbs and genotyping. CONCLUSIONS: Typing of human enteroviruses can be effectively performed using an integration of antibody-based and molecular methods.

Molecular typing of clinical adenovirus specimens by an algorithm which permits detection of adenovirus coinfections and intermediate adenovirus strains.

BACKGROUND: Epidemiological data suggest that clinical outcomes of human adenovirus (HAdV) infection may be influenced by virus serotype, coinfection with multiple strains, or infection with novel intermediate strains. In this report, we propose a clinical algorithm for detecting HAdV coinfection and intermediate strains. STUDY DESIGN: We PCR amplified and sequenced subregions of the hexon and fiber genes of 342 HAdV-positive clinical specimens obtained from 14 surveillance laboratories. Sequences were then compared with those from 52 HAdV prototypic strains. HAdV-positive specimens that showed nucleotide sequence identity with a corresponding prototype strain were designated as being of that strain. When hexon and fiber gene sequences disagreed, or sequence identity was low, the specimens were further characterized by viral culture, plaque purification, repeat PCR with sequencing, and genome restriction enzyme digest analysis. RESULTS: Of the 342 HAdV-positive clinical specimens, 328 (95.9%) were single HAdV strain infections, 12 (3.5%) were coinfections, and 2 (0.6%) had intermediate strains. Coinfected specimens and intermediate HAdV strains considered together were more likely to be associated with severe illness compared to other HAdV-positive specimens (OR=3.8; 95% CI=1.2-11.9). CONCLUSIONS: The majority of severe cases of HAdV illness cases occurred among immunocompromised patients. The analytic algorithm we describe here can be used to screen clinical specimens for evidence of HAdV coinfection and novel intermediate HAdV strains. This algorithm may be especially useful in investigating HAdV outbreaks and clusters of unusually severe HAdV disease.

Emergent US adenovirus 3 strains associated with an epidemic and serious disease.

BACKGROUND: Adenovirus type 3 (HAdV3) is one of the most prevalent serotypes detected globally. Variants of HAdV3 have been associated with outbreaks of severe disease. OBJECTIVES: To better understand genetic diversity of circulating HAdV3s and examine risk factors for severe disease. STUDY DESIGN: Restriction enzyme analysis for genomic characterization of clinical HAdV3 isolates detected by 15 collaborative US laboratories during the period July 2004 to May 2007. Multivariate modeling was employed for statistical analyses. RESULTS: The most common HAdV3 types of 516 isolates studied were HAdV3a2 (36.9%), HAdV3a50 (27.1%), HAdV3a51 (18.0%), and HAdV3a17 (4.6%). Non-HAdV3a genome types were rare (1.2%). HAdV3a50 and HAdV3a51 are newly described variants which became more prevalent in 2006 and 2007 and have been associated with at least one epidemic. Their uniqueness was determined by specific banding profiles generated by digests with endonucleases BclI, BglII, and HindIII. Multivariable risk factor modeling demonstrated that children under 2 years of age (OR=2.7; 95%CI 1.6-4.6), persons with chronic disease (OR=5.1; 95%CI 2.6-9.8), persons infected with HAdV3a2 (OR=3.0; 95%CI 1.5-6.0), with HAdV3a50 (OR=2.5; 95%CI 1.2-5.2), or with multiple or rare strains (OR=2.8; 95%CI 1.3-6.5) were at increased risk of severe HAdV3 clinical disease. CONCLUSIONS: In the study period considerable genetic diversity was found among US clinical HAdV3 strains. Novel variants emerged and became prevalent. One such emergent strain may be associated with more severe clinical disease.

Analysis of a new human parechovirus allows the definition of parechovirus types and the identification of RNA structural domains.

Human parechoviruses (HPeV), members of the Parechovirus genus of Picornaviridae, are frequent pathogens but have been comparatively poorly studied, and little is known of their diversity, evolution, and molecular biology. To increase the amount of information available, we have analyzed 7 HPeV strains isolated in California between 1973 and 1992. We found that, on the basis of VP1 sequences, these fall into two genetic groups, one of which has not been previously observed, bringing the number of known groups to five. While these correlate partly with the three known serotypes, two members of the HPeV2 serotype belong to different genetic groups. In view of the growing importance of molecular techniques in diagnosis, we suggest that genotype is an important criterion for identifying viruses, and we propose that the genetic groups we have defined should be termed human parechovirus types 1 to 5. Complete nucleotide sequence analysis of two of the Californian isolates, representing two types, confirmed the identification of a new genetic group and suggested a role for recombination in parechovirus evolution. It also allowed the identification of a putative HPeV1 cis-acting replication element, which is located in the VP0 coding region, as well as the refinement of previously predicted 5' and 3' untranslated region structures. Thus, the results have significantly improved our understanding of these common pathogens.