ImportanceSARS-CoV-2 infection is associated with persistent, relapsing, or new symptoms or other health effects occurring after acute infection, termed postacute sequelae of SARS-CoV-2 infection (PASC), also known as long COVID. Characterizing PASC requires analysis of prospectively and uniformly collected data from diverse uninfected and infected individuals.ObjectiveTo develop a definition of PASC using self-reported symptoms and describe PASC frequencies across cohorts, vaccination status, and number of infections.Design, Setting, and ParticipantsProspective observational cohort study of adults with and without SARS-CoV-2 infection at 85 enrolling sites (hospitals, health centers, community organizations) located in 33 states plus Washington, DC, and Puerto Rico. Participants who were enrolled in the RECOVER adult cohort before April 10, 2023, completed a symptom survey 6 months or more after acute symptom onset or test date. Selection included population-based, volunteer, and convenience sampling.ExposureSARS-CoV-2 infection.Main Outcomes and MeasuresPASC and 44 participant-reported symptoms (with severity thresholds).ResultsA total of 9764 participants (89% SARS-CoV-2 infected; 71% female; 16% Hispanic/Latino; 15% non-Hispanic Black; median age, 47 years [IQR, 35-60]) met selection criteria. Adjusted odds ratios were 1.5 or greater (infected vs uninfected participants) for 37 symptoms. Symptoms contributing to PASC score included postexertional malaise, fatigue, brain fog, dizziness, gastrointestinal symptoms, palpitations, changes in sexual desire or capacity, loss of or change in smell or taste, thirst, chronic cough, chest pain, and abnormal movements. Among 2231 participants first infected on or after December 1, 2021, and enrolled within 30 days of infection, 224 (10% [95% CI, 8.8%-11%]) were PASC positive at 6 months.Conclusions and RelevanceA definition of PASC was developed based on symptoms in a prospective cohort study. As a first step to providing a framework for other investigations, iterative refinement that further incorporates other clinical features is needed to support actionable definitions of PASC.

Background: Haemophilus influenzae requires heme for aerobic growth and possesses multiple mechanisms to obtain this essential nutrient. Although an understanding of the heme acquisition mechanisms of H. influenzae is emerging, significant gaps in our knowledge remain. Unresolved issues include the identities of all genes exhibiting altered transcription in response to iron and heme availability, the fraction of such genes functioning in iron/heme acquisition, and the heterogeneity of this gene set among clinical isolates. Previously we utilized H. influenzae strain Rd KW20 to demonstrate the utility of transcriptional profiling in defining the genes exhibiting altered transcription in response to environmental iron and heme levels. The current study expands upon those observations by determining the iron/heme modulons of two clinical isolates, the type b isolate 10810 and the nontypeable isolate R2866. These data are used to begin to define the core iron/heme modulon of the species.

Haemophilus influenzae has an absolute growth requirement for a porphyrin source. This growth requirement can be satisfied in vitro by haem, haemoglobin or the haemoglobin-haptoglobin, haem-haemopexin and haem-albumin complexes. A family of proteins, termed the Hgp proteins, which are essential for utilization of the haemoglobin-haptoglobin complex, has previously been identified. A strain lacking the Hgp proteins also has a residual ability to utilize haemoglobin, indicating that additional moieties contribute to haemoglobin utilization. Using a haemoglobin affinity method an approximately 105 kDa protein was isolated. Mutation of the identified gene in an Hgp null background reduced the ability of the mutant strain to utilize haemoglobin in vitro. The mutation also resulted in a reduced ability to utilize haem, haem-haemopexin, haem-albumin and haemoglobin-haptoglobin, thus identifying a general haem-utilization protein (Hup) in Haemophilus influenzae.

Haemophilus influenzae has an absolute growth requirement for heme and a heme binding lipoprotein (HbpA) has been implicated in the utilization of this essential nutrient. HbpA was identified by examining clones from an H. influenzae genomic library that caused Escherichia coli harboring the clone to bind heme. However, HbpA has not been shown to mediate heme acquisition in H. influenzae. We constructed an insertional mutation of hbpA in a nontypeable H. influenzae strain and demonstrated a role for the gene in utilization of multiple heme sources. This is the first report confirming a role for HbpA in utilization of heme.

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Haemophilus influenzae has an absolute growth requirement for a porphyrin source, which can be supplied in vitro by haem, haemoglobin, or the haemoglobin-haptoglobin, haem-haemopexin and haem-albumin complexes. Utilization of the haem-haemopexin complex is known to be mediated by the products of the hxuCBA gene cluster. It was demonstrated that hxuC, but not hxuA or hxuB, is also essential for the utilization of haem from haem-albumin complexes. Mutants of the type b strain E1a lacking genes in the hxuCBA gene cluster were examined for their ability to cause bacteraemia in rat models of invasive disease. In 5-day-old rats, mutants in the hxuCBA genes yielded a significantly reduced bacteraemic titre compared to the wild-type strain. In addition, 5-day-old rats infected with the hxuCBA mutant strains exhibited significantly improved survival rates compared to those infected with the wild-type strain. Mutations in the haemoglobin/haemoglobinhaptoglobin-binding protein genes (hgps), either alone or in combination with the hxuCBA mutations, had no impact on virulence in 5-day-old rats. In 30-day-old rats infected with either the hxuCBA mutants or the wild-type strains, there was no significant difference in the ability to establish bacteraemia although bacterial titres were lower in rats infected with the hxuCBA mutants than in those infected with the wild-type strain. These age-related differences in the impact of mutations in the hxuCBA gene cluster may be related to changes in levels of host haem-binding proteins during development of the rat.

Haemophilus influenzae requires either heme or a porphyrin and iron source for growth. Microarray studies of H. influenzae strain Rd KW20 identified 162 iron/heme-regulated genes, representing ϳ10% of the genome, with >1.5-fold changes in transcription in response to iron/heme availability in vitro. Eighty genes were preferentially expressed under iron/heme restriction; 82 genes were preferentially expressed under iron/hemereplete conditions.

Burkholderia cenocepacia is a member of the Burkholderia cepacia complex, a group of genetically similar species that inhabit a number of environmental niches, including the lungs of patients with cystic fibrosis (CF). To colonize the lung, this bacterium requires a source of iron to satisfy its nutritional requirements for this important metal. Because of the high potential for damage in lung tissue resulting from oxygen–iron interactions, this metal is sequestered by a number of mechanisms that render it potentially unavailable to invading micro-organisms. Such mechanisms include the intracellular and extracellular presence of the iron-binding protein ferritin. Ferritin has a highly stable macromolecular structure and may contain up to 4500 iron atoms per molecule. To date, there has been no known report of a pathogenic bacterial species that directly utilizes iron sequestered by this macromolecule. To examine the ability of ferritin to support growth of B. cenocepacia J2315, iron-deficient media were supplemented with different concentrations of ferritin and the growth kinetics characterized over a 40 h period. The results indicated that B. cenocepacia J2315 utilizes iron bound by ferritin. Further studies examining the mechanisms of iron uptake from ferritin indicated that iron utilization results from a proteolytic degradation of this otherwise stable macromolecular structure. Since it is known that the ferritin concentration is significantly higher in the CF lung than in healthy lungs, this novel iron-acquisition mechanism may contribute to infection by B. cenocepacia in people with CF.