Metabolic Analysis of Moraxella catarrhalis and the Effect of Selected In Vitro Growth Conditions on Global Gene Expression

Abstract

The nucleotide sequence from the genome of Moraxella catarrhalis ATCC 43617 was annotated and used both to assess the metabolic capabilities and limitations of this bacterium and to design probes for a DNA microarray. An absence of gene products for utilization of exogenous carbohydrates was noteworthy and could be correlated with published phenotypic data. Gene products necessary for aerobic energy generation were present, as were a few gene products generally ascribed to anaerobic systems. Enzymes for synthesis of all amino acids except proline and arginine were present. M. catarrhalis DNA microarrays containing 70-mer oligonucleotide probes were designed from the genome-derived nucleotide sequence data. Analysis of total RNA extracted from M. catarrhalis ATCC 43617 cells grown under iron-replete and iron-restricted conditions was used to establish the utility of these DNA microarrays. These DNA microarrays were then used to analyze total RNA from M. catarrhalis cells grown in a continuous-flow biofilm system and in the planktonic state. The genes whose expression was most dramatically increased by growth in the biofilm state included those encoding a nitrate reductase, a nitrite reductase, and a nitric oxide reductase. Real-time reverse transcriptase PCR analysis was used to validate these DNA microarray results. These results indicate that growth of M. catarrhalis in a biofilm results in increased expression of gene products which can function not only in energy generation but also in resisting certain elements of the innate immune response.Moraxella catarrhalis is a gram-negative, unencapsulated bacterium that can colonize the mucosal surface of the human nasopharynx, most frequently in infants and very young children (22). When this organism traverses the eustachian tube in these very young individuals, it can cause otitis media (7). Alternatively, in colonized adults, M. catarrhalis gains access to the bronchi and there causes exacerbations of chronic obstructive pulmonary disease (48). This organism can also infrequently cause other types of infections (for reviews, see references 39 and 73).Information about the virulence mechanisms employed by this organism in the production of disease is still very limited, although a number of putative virulence factors have been identified, including proteins located in or attached to the outer membrane (1, 6, 9, 23, 24, 26, 32-34, 40, 44, 49, 51, 54, 57, 65, 71), as well as lipooligosaccharide (42, 59). Validation of the actual involvement of these different gene products in disease processes has been severely hindered by the lack of an appropriate animal model for M. catarrhalis disease (39). Similarly, little is known about how M. catarrhalis colonizes the nasopharynx, and while several M. catarrhalis adhesins which function in vitro have been identified (33,40,61,71), the relative importance of these macromolecules in the colonization process in vivo remains to be determined. Recent studies aimed at addressing this issue have included the use of reverse transcr...

Molecular Aspects ofMoraxella catarrhalisPathogenesis

“…For instance, M. catarrhalis harbors an intact glyoxylate pathway, which is consistent with the ability to utilize acetate for energy generation. In addition, all enzymes of the gluconeogenic pathway are present, indicating the importance of this pathway (152). Unfortunately, the relationship between virulence and in vivo nutrient acquisition is a less-studied aspect of M. catarrhalis pathogenesis, although a few mechanisms have been shown to be particularly important for nutrient acquisition.…”

“…Typically, M. catarrhalis is not able to utilize exogenous carbohydrates, because it does not harbor the enzymes necessary for transport and processing of these compounds, i.e., it harbors an incomplete Embden-Meyerhof-Parnas pathway, it has an incomplete pentose cycle, and enzymes of the Entner-Dourdoroff pathway are completely absent (152). M. catarrhalis can utilize acetate, lactate, and fatty acids for growth and is auxotrophic for arginine and possibly proline (78,152). Other pathogens that colonize the nasopharynx, such as N. meningitidis and H. influenzae, are also able to utilize only a limited number of carbohydrates, suggesting that the nasopharynx offers a limited carbohydrate availability or a limited variety of carbohydrates (29,152).…”

“…Unfortunately, the relationship between virulence and in vivo nutrient acquisition is a less-studied aspect of M. catarrhalis pathogenesis, although a few mechanisms have been shown to be particularly important for nutrient acquisition. These include iron acquisition (119), the nitrate respiratory chain (152), and the M35 porin, which appears to be essential for growth under nutrient-poor conditions (36).…”

“…For example, M. catarrhalis cannot grow under strictly anaerobic conditions (5), but enzymes involved in anaerobic metabolism are present, and they could confer the ability to grow under microaerophilic conditions (152). Typically, M. catarrhalis is not able to utilize exogenous carbohydrates, because it does not harbor the enzymes necessary for transport and processing of these compounds, i.e., it harbors an incomplete Embden-Meyerhof-Parnas pathway, it has an incomplete pentose cycle, and enzymes of the Entner-Dourdoroff pathway are completely absent (152). M. catarrhalis can utilize acetate, lactate, and fatty acids for growth and is auxotrophic for arginine and possibly proline (78,152).…”

“…M. catarrhalis can utilize acetate, lactate, and fatty acids for growth and is auxotrophic for arginine and possibly proline (78,152). Other pathogens that colonize the nasopharynx, such as N. meningitidis and H. influenzae, are also able to utilize only a limited number of carbohydrates, suggesting that the nasopharynx offers a limited carbohydrate availability or a limited variety of carbohydrates (29,152). This means that other energy-yielding biochemical pathways are likely to be especially important to M. catarrhalis.…”