Invisible Alliances: How Bacteria Communicate, Cooperate and Cause Infections

Exploring the biological properties of bacterial monocultures and their co-cultured variations in Enterobacter, Citrobacter, Serratia, and Proteus species.

Introduction

Beyond the world visible to the naked eye exists an invisible universe of microorganisms. Bacteria of the genera Enterobacter, Citrobacter, Serratia, and Proteus — typical representatives of the Enterobacteriaceae family — are most often mentioned in the context of infectious diseases. However, these microscopic organisms lead a much more complex and organized life than might appear at first glance.

When they exist alone (as monocultures), their properties are well studied. But the real mysteries begin when different bacterial species find themselves together in one environment — their coexistence radically changes the biological properties, behavior, and pathogenicity of each participant in this invisible alliance.

The Microscopic World: Who's Who in the Bacterial Community

Enterobacteriaceae: The Large Bacterial Family

The Enterobacteriaceae family represents gram-negative rod-shaped bacteria that are widely distributed in the environment and are often part of the normal intestinal flora of animals and humans 1 . They belong to the class Gammaproteobacteria and include more than 68 genera with 355 species 1 .

These bacteria are facultative anaerobes (can live both in the presence and absence of oxygen), do not form spores, and usually have flagella for movement, with the exception of some species of Klebsiella and Shigella 1 .

Individual Portraits of Our "Heroes"

Each genus of bacteria has unique characteristics that determine its behavior both in monoculture and in mixed communities.

  • Proteus spp. - known for their "swarming" phenomenon 4 7
  • Serratia marcescens - known biofilm former 3
  • Citrobacter spp. - demonstrates tropism for the nervous system 9
  • Enterobacter spp. - often cause nosocomial infections 5

Characteristic Biological Properties of Bacteria in Monoculture

Bacterial Genus Motility Biofilm Formation Key Pathogenic Properties
Proteus Active "swarming" Moderate ability Urease activity, kidney stone formation
Serratia Motile Intensive formation of filamentous biofilms Quorum sensing regulation, antibiotic resistance
Citrobacter Motile Capable of biofilm formation Tropism for nervous system, intracellular survival
Enterobacter Motile Capable of biofilm formation Nosocomial infections, antibiotic resistance

Co-Culturing Phenomenon: When the Whole is Greater Than the Sum of Its Parts

When different bacterial species find themselves together in one environment, their properties can change radically. This occurs thanks to various mechanisms of intermicrobial interaction:

Quorum Sensing System

The ability of bacteria to coordinate behavior through chemical signals (acylated homoserine lactones), which plays a key role in the formation of mixed biofilms 3 .

Horizontal Gene Transfer

Exchange of antibiotic resistance and virulence genes between different bacterial species 1 6 .

Metabolic Cooperation

When metabolic products of some bacteria serve as nutrients for others 2 .

Physical Interaction

For example, the ability of Proteus mirabilis to move non-motile bacteria such as Klebsiella pneumoniae along catheter surfaces 7 .

Experiment: How Citrobacter Increases the Energy Value of Food in Community with Other Bacteria

Methodology

One illustrative example of the influence of co-culturing on the biological properties of bacteria is a study that examined the role of Citrobacter in increasing energy storage in fish on a high-fat diet 2 .

Experimental Steps:
  1. Isolation of Citrobacter strain (designated S1) from the intestine of Nile tilapia using media with soybean oil as the main carbon source.
  2. Determination of the 16S rRNA gene sequence for accurate strain identification.
  3. Addition of strain S1 to fish diet at a concentration of 10⁹ CFU/g feed.
  4. Conducting four experimental groups: control (basal diet), control + S1, high-fat diet, high-fat diet + S1.
  5. Assessment of intestinal colonization by S1 bacteria by fecal culture on selective media.
  6. Measurement of fish growth, body lipid content, and Citrobacter population level using PCR with species-specific primers.
  7. Analysis of intestinal microbial composition by sequencing the V3-V4 region of the 16S rRNA gene.
Experimental Design

Results and Analysis

The study demonstrated several important effects of co-culturing:

Colonization and Growth

The Citrobacter S1 strain successfully colonized the fish intestine, with its numbers significantly increasing in the high-fat diet group 2 .

Impact on Metabolism

Addition of S1 bacteria to the high-fat diet significantly increased body lipid content in fish compared to all other groups 2 . This was accompanied by increased triglyceride absorption efficiency and intestinal permeability.

This research demonstrates that even non-dominant bacteria in the intestine, such as Citrobacter, can significantly influence host physiology, especially in conditions of coexistence with other members of the microbiome 2 .

Influence of Citrobacter and High-Fat Diet on Fish Parameters
Parameter Control Control + S1 High-Fat Diet High-Fat Diet + S1
Body Lipid Content Low Low Moderate Significantly Increased
Triglyceride Absorption Efficiency Basal level Slight increase Moderate increase Significant increase
Citrobacter Count in Intestine Low Moderate Moderate High
Relative Firmicutes Content 5.11% 23.98% 58.15% 62.64%

Antibiotic Resistance and Biofilms: A Challenge for Modern Medicine

Biofilms as a Survival Strategy

Biofilms are complex communities of microorganisms enclosed in a matrix of extracellular polymers synthesized by the microorganisms themselves 6 . This matrix provides protection from adverse environmental conditions, antibiotic actions, and the host's immune system .

A 2020 study analyzing Enterobacteriaceae strains from meat products showed that out of 200 strains, 46 (23.0%) were strong biofilm producers, 60 (30.0%) were moderate, and 79 (39.5%) were weak 6 .

Antibiotic Resistance

The problem of antibiotic resistance among Enterobacteriaceae has reached alarming proportions. A 2020 study demonstrated that 63% of Enterobacteriaceae strains isolated from meat products were multidrug-resistant (resistant to two or more antibiotics) 6 . The greatest resistance was observed to ampicillin, cefotaxime, ceftazidime, and streptomycin 6 .

Antibiotic Resistance Among Enterobacteriaceae (Data from 200 Strains) 6

Antibiotic Percentage of Resistant Strains Medical Significance
Ampicillin 65.5% Critically important antimicrobial
Cefotaxime 54.5% 3rd generation cephalosporin
Ceftazidime 52.0% 3rd generation cephalosporin
Streptomycin 50.0% Aminoglycoside
Tetracycline 35.5% Broad spectrum
Nalidixic Acid 30.0% Quinolone
Gentamicin 18.5% Aminoglycoside
Ciprofloxacin 16.5% Fluoroquinolone
Antibiotic Resistance Visualization

Scientist's Toolkit: Methods for Studying Bacterial Interactions

Modern research on bacterial co-culturing requires the use of various methods and reagents:

Confocal and Fluorescence Microscopy

Allows visualization of biofilm structure and arrangement of different bacterial species within them 8 .

Molecular Docking

Computer modeling of interactions between potential antibacterial compounds and targets in bacterial cells .

Next-Generation Sequencing

Determination of the complete genomic composition of microbial communities 8 .

Flow Cytometry

Assessment of bacterial viability and physiological state .

Spectrofluorimeters

Instruments for measuring fluorescence, used to study molecular interactions 8 .

Conclusion

The study of biological properties of bacterial monocultures and their co-cultured variations opens new horizons in understanding the microbial world. It turns out that bacteria are not solitary players, but participants in complex communities where interaction between different species can radically change their properties, pathogenicity, and treatment resistance.

Understanding these mechanisms is necessary for developing new approaches in combating infectious diseases, especially in an era of growing antibiotic resistance. Perhaps the key to solving the problem is not destroying bacteria, but managing their interactions, disrupting communication between them, and targeted impact on mechanisms of collective behavior.

Invisible alliances of bacteria continue to be one of the most fascinating mysteries of microbiology, the unraveling of which could lead to revolutionary discoveries in medicine and biotechnology.

References