Acute respiratory infections are responsible for an estimated 4 million deaths annually worldwide, and are the leading cause of death in children younger than 5 years. Over 1 million people in the United States are hospitalized each year with pneumonia. Mycobacterium tuberculosis infects one third of world's population. There are more than 1 million tuberculosis-related deaths worldwide each year. Emerging resistance to multiple available antimicrobial agents has hampered the ability to treat tuberculosis and hospital-acquired respiratory infections. The laboratory diagnosis of respiratory infections is an important part of patient management and treatment. In addition to culture isolation of pathogens, advances have been made in a number of non-culture methods. This issue of Clinics in Laboratory Medicinee reviews state-of-the-art laboratory diagnosis of respiratory infections, as well as the testing of susceptibility to antibiotics and antiviral agents. Among some of the respiratory infections covered are: Cystic fibrosis infections; Pertussis; Pharyngitis; Fungal infections. Among the diagnostic tests are: Interferon gamma release assays; Molecuar diagnosis of TB; Urine antigen tests and discussion of Antibiotic resistance in nosocomial respiratory infections.
Front Cover 1
Respiratory Infections 2
copyright
3
Contributors 4
Contents 6
Clinics In Laboratory Medicine
10
Preface
12
Infections in Patients with Cystic Fibrosis 14
Key points 14
Introduction, epidemiology, and clinical presentation 14
Microbiology, diagnostic considerations, and susceptibility testing of agents of chronic infection 16
Staphylococcus aureus 16
Pseudomonas aeruginosa 20
Burkholderia cepacia complex and related organisms 22
Stenotrophomonas and Achromobacter 24
Mycobacterium 25
Fungi 26
Respiratory viruses 27
Microbiome considerations 27
References 28
Urine Antigen Tests for the Diagnosis of Respiratory Infections 36
Key points 36
Introduction 36
Microbiology 37
Legionella 37
Pneumococcus 37
Histoplasma 37
Epidemiology, disease presentation, and pathogenesis 38
Legionella 38
Pneumococcus 38
Histoplasma 39
Diagnosis 39
Legionella Urinary Antigen Testing 40
ELISA 40
LFA 40
Pneumococcal Urinary Antigen Test 42
Effects of study design on the sensitivity of the pUAT 42
Colonization and the pUAT 44
Histoplasma Antigen Testing 44
Miravista Diagnostics enzyme immunoassay 44
Second generation 44
Third generation 45
Immuno-Mycologics EIA 45
Treatment 46
Empiric Therapy for LD/Pneumococcal Pneumonia 46
Targeted Therapy for LD/Pneumococcal Pneumonia 46
Treatment of Histoplasma 46
Summary and discussion 47
References 47
Pertussis 54
Key points 54
Introduction 54
Microbiology 55
Epidemiology 55
Clinical presentations 57
Pathogenesis, immunity, and vaccination 57
Diagnosis 59
Case definitions 59
Laboratory testing 60
Specimens 60
Test Methods 60
Direct fluorescent antibody 62
Culture 62
Nucleic acid amplification tests 62
Pretreatment and extraction 62
Targets for PCR 63
Low copy number and high CT values 64
NAATs other than PCR 64
Contamination 65
Interpretation 65
Serology 65
Treatment 66
Summary/discussion 66
References 67
Antibiotic Resistance in Nosocomial Respiratory Infections 74
Key points 74
Introduction 74
Etiology 75
Epidemiology 76
Clinical presentation 77
Pathogenesis 77
Diagnosis 78
Treatment 83
Summary 84
References 84
Nontuberculous Mycobacteria in Respiratory Infections 88
Key points 88
Microbiology 89
Epidemiology 89
Pathogenesis and clinical significance 92
Diagnosis 94
Specimen processing 95
Acid-fast microscopy 96
Direct NAA assays 97
Growth detection 97
Identification 99
Nucleic Acid Hybridization Methods 99
PCR and Restriction Fragment Length Polymorphism Analysis 100
Line Probe Assays 101
DNA Sequencing 101
MALDI-TOF MS 102
Genotyping 103
Antimicrobial susceptibility testing 103
Identified focus areas in NTM research 105
References 106
Molecular Diagnosis of Tuberculosis and Drug Resistance 114
Key points 114
Introduction 114
Microbiology 115
Epidemiology 115
Clinical presentation 115
Pathogenesis of pulmonary tuberculosis 116
Molecular diagnosis 116
Genes Associated with Drug Resistance 116
Isoniazid 118
Rifampin and rifabutin 119
Ethambutol 120
Pyrazinamide 120
Fluoroquinolones 120
Injectable drugs 121
Molecular assays 121
Molecular Beacon Assays 121
Line-Probe Assays 124
Sanger Sequencing 124
Pyrosequencing 124
Next-Generation Sequencing 125
Practical use of molecular diagnostic assays 125
Impact on patient management, TB control, and infection control 125
Summary 126
References 127
Nonmolecular Methods for the Diagnosis of Respiratory Fungal Infections 132
Key points 132
Introduction 132
Microbiology/Epidemiology 133
Clinical presentation 134
Pathogenesis 135
Diagnosis 135
Invasive Aspergillosis (IA) 136
GM assay 136
BG assay 139
Aspergillus lateral-flow devices 140
Anti-Aspergillus antibodies 140
Chronic Pulmonary Aspergillosis 141
Pneumonia Caused by Non-Aspergillus Molds 141
Pneumocystis jirovecii Pneumonia 141
Pulmonary Cryptococcosis 142
Treatment 143
Discussion/Summary 143
References 145
Interferon-Gamma Release Assays 154
Key points 154
Introduction 154
Microbiology 155
Epidemiology 155
Clinical presentation 156
Pathogenesis 157
Diagnosis 158
Interferon-Gamma Release Assays 158
Test Performance of IGRAs 159
IGRAs in Targeted Populations 160
Treatment 162
Summary/Discussion 162
References 163
Respiratory Fungal Infections 168
Key points 168
Introduction 168
Aspergillosis 170
Diagnosis 170
Combination Testing 171
The Need for Prospective Clinical Trials 172
Pneumocystis jirovecii pneumonia 173
Introduction 173
Microbiology 173
Epidemiology 173
Clinical Presentation 174
Pathogenesis 174
Laboratory Diagnosis 174
PCR or Antigen Tests 176
Treatment 176
Summary 176
References 177
Rapid Diagnosis of Influenza 182
Key points 182
Influenza viruses 182
Pathogenesis 183
Clinical Presentation 183
Treatment 183
General principles of laboratory diagnosis of influenza infection 184
Sample collection and transport 186
Diagnostic methods 187
Viral Culture 187
Viral Antigen Detection 187
Immunofluorescence 187
Lateral flow IC 188
Performance of RIDTs 188
Nucleic Acid Detection 190
Conventional PCR 190
Real-time PCR 191
Multiplex methods 192
Rapid NAAT for the detection of influenza viruses 192
Description of the Systems 193
Assay Performance 195
Limitations and Future Developments 196
Factors to consider 196
Summary 198
References 198
Antiviral Resistance in Influenza Viruses 204
Key points 204
Introduction 204
Microbiology 205
Epidemiology and transmission 206
Pathogenesis 206
Clinical presentation 206
Pharmacologic agents 207
Adamantanes 207
NA Inhibitors 207
Other Antiviral Targets and Drugs 208
Resistance genetics 210
Influenza diagnosis 211
Antiviral drug susceptibility testing 212
Genotypic Methods 212
Phenotypic Methods 214
Patient Testing and Surveillance Programs 215
Treatment and prognosis 216
Summary 217
Acknowledgments 217
References 217
Emerging Respiratory Viruses Other than Influenza 226
Key points 226
Introduction 226
Microbiology 227
MERS-CoV 227
Ad14 228
RV-C 229
HBoV1 229
Epidemiology 230
MERS-CoV 230
Ad14 230
RV-C 231
HBoV1 232
Clinical presentation 232
MERS-CoV 232
Ad14 232
RV-C 233
HBoV1 233
Pathogenesis 234
MERS-CoV 234
Ad14 234
RV-C 234
HBoV1 236
Diagnosis 237
Treatment and prognosis 238
Summary 239
References 240
Index 248
Infections in Patients with Cystic Fibrosis
Diagnostic Microbiology Update
Peter H. Gilligan, PhD, DABMMgilliganncphd@gmail.com, Pathology-Laboratory Medicine and Microbiology-Immunology, Clinical Microbiology-Immunology Laboratories, UNC Health Care, UNC Hospitals, UNC School Medicine, Room 1035, CB 7600, Chapel Hill, NC 27516, USA
Survival has improved in patients with cystic fibrosis (CF), in part because of aggressive antimicrobial management. Two multidrug-resistant environmental bacteria, the Burkholderia cepacia group and nontuberculous mycobacteria, have emerged. Improving genomic and proteomic technologies are allowing better identification of bacteria and fungi found in the CF lung and detection of viral agents that may be associated with pulmonary exacerbations. Anaerobic bacteria and Streptococcus angionsus group organisms may play a role in chronic CF lung infections. The diversity of organisms declines perhaps as a result of aggressive antimicrobial therapy, and an apex predator, Pseudomonas aeruginosa, may emerge in many patients with CF.
Keywords
Infection
Cystic fibrosis
Diagnostic microbiology
Update
Key points
• Cystic fibrosis is the most important genetic disease in Caucasians. Patients with this disease die prematurely primarily as a result of chronic lung infection. Staphylococcus aureus and mucoid Pseudomonas aeruginosa continue to be the key pulmonary pathogens.
• Survival has improved in patients with CF in part because of aggressive antimicrobial management. An unintended consequence of this therapy has been the emergence of 2 multidrug-resistant environmental bacteria: the Burkholderia cepacia group and nontuberculous mycobacteria.
• Burkholderia cenocepacia, a species within the Burkholderia cepacia complex, is associated with high mortality and is a contraindication for lung transplantation. The key nontuberculous mycobacterial pathogen, Mycobacterium abscessus, is not so virulent and is not a lung transplantation contraindication. Both present an infection control challenge, because they can be spread from person to person.
• Improving genomic and proteomic technologies are allowing better identification of bacteria and fungi found in the CF lung and to detect viral agents that may be associated with pulmonary exacerbations. Chronic rhinovirus infections are of particular interest.
• Microbiome studies have identified 2 groups of bacteria that may play a role in chronic CF lung infections: anaerobic bacteria and Streptococcus angionsus group organisms. Microbiome studies also show that as the diversity of organisms decline, perhaps as a result of aggressive antimicrobial therapy, an apex predator, etc., Pseudomonas aeruginosa, may emerge in many patients with CF.
Introduction, epidemiology, and clinical presentation
Cystic fibrosis (CF) is the most common autosomal-recessive genetic disease that occurs in non-Hispanic Caucasians populations, although other racial groups may have this disease as well.1 Affected individuals have mutation in the CF transmembrane conductance gene (CFTR), a membrane protein involved in sodium and chloride transport in epithelial cells.2 The resulting dysregulation in electrolyte transport leads to depletion in airway surface liquid on bronchial epithelial cell surfaces. As a result, patients with CF have thick, dry, tenacious mucus, which impairs mucociliary clearance of particulates, especially bacteria and fungal conidia, from the airways. This environment is ideal for the growth of a limited number of organisms, primarily those that thrive in natural environments such as water. This thickened mucus provides an ideal niche for the establishment of chronic infection. It is this chronic infection that results in the premature death that in seen in CF.3
More than 1800 CFTR mutations have been associated with CF.4 The most common mutation is F508del, which is found in ∼85% of people in the United States; approximately 47% are homozygous for this mutated gene.4 Further carrier rate for mutated CFTR genes is estimated to range from 1/25 for non-Hispanic Caucasians to 1/61 for African Americans to 1/94 for Asian Americans.1 CF is seen most frequently in North America, Northern Europe, Australia, New Zealand, Brazil, and Argentina. It is estimated that 1 in 3500 live births result in clinical disease.4
Currently life expectancy in US patients with CF is approximately 38 years, significantly less than that of the general population.4 Cardiopulmonary failure secondary to chronic lung disease is responsible for 85% of premature deaths in CF. The airways of patients with CF become infected in infancy. This situation begins periods of chronic infection and lung inflammation with accompanying cough, which is a lifelong reality in patients with CF. A hallmark of chronic infection and airway inflammation is periods of pulmonary exacerbations. Pulmonary exacerbations are characterized by worsening symptoms, including increased cough and sputum production, hemoptysis, shortness of breath, increased respiratory rate, loss of appetite, weight loss, increased neutrophil counts, and declining pulmonary function.5 The events that trigger these pulmonary exacerbations are not clearly understood, although viral infections and perhaps changes in the microbiome may be important.6,7 Exacerbations are characterized by the recruitment of neutrophils, cytokine release, and high level of neutrophil-derived elastases in the bronchi and bronchioles, causing significant lung disease.8 Antimicrobial therapy has been shown to be effective in treating exacerbations symptomatically.9 However, over time, lung function deteriorates and becomes so low, that it is no longer compatible with life (Fig. 1). Only lung transplantation can successfully reverse this disease course.8
Fig. 1 Lung function by age group, 2011. FEV1, forced expiratory volume in 1 second. (From Cystic Fibrosis Foundation. Cystic Fibrosis Foundation patient registry 2011 annual data report. 2012.)
Microbiology, diagnostic considerations, and susceptibility testing of agents of chronic infection
Over the past 4 decades, our understanding of the complex nature of chronic lung infections has greatly expanded. Over the past 3 decades, there has been more than a doubling in life expectancy in the population with CF.4 Three factors have been central to this improvement:
a. More effective antimicrobial therapy and treatment strategies, with early eradication of Pseudomonas aeruginosa being a key strategy
b. Improvement in airway clearance techniques
c. Improvements in infection control techniques to prevent the spread of organisms highly virulent to patients with CF, especially Burkholderia cenocepacia2,9
With the use of broader-spectrum antimicrobials, we are seeing a plethora of emerging highly resistant bacteria and fungi in the CF airways. Our understanding of the role of these organisms in chronic infection and inflammation is poorly delineated (Box 1). Over the past decades, new technologies (Fig. 2) have been developed and applied to this understanding. These technologies include nucleic acid amplification techniques (NAATs) for direct organism detection, including multiplex NAAT for viruses; the use of DNA sequence analysis for organism identification; molecularly based epidemiologic techniques, including pulsed field gel electrophoresis (PFGE), multilocus sequence type, whole genome sequencing; and matrix-assisted laser desorption ionization–time of flight mass spectroscopy (MALDI-TOF MS).
Box 1 Pathogenic potential of commonly recovered organisms from chronic CF airway infections or pulmonary exacerbations
• Known
Pseudomonas aeruginosa
Staphylococcus aureus
Methicillin resistant
Small colony variant
Burkholderia multivorans
Burkholderia cenocepacia
Burkholderia dolosa
Aspergillus spp
Scedosporium spp
Mycobacterium abscessus
Influenza virus
Respiratory syncytial virus
• Possible/likely
Haemophilus influenzae
Mycobacterium avium complex
Anaerobic bacteria especially Prevotella...
| Erscheint lt. Verlag | 9.8.2014 |
|---|---|
| Sprache | englisch |
| Themenwelt | Medizinische Fachgebiete ► Innere Medizin ► Pneumologie |
| ISBN-10 | 0-323-29942-3 / 0323299423 |
| ISBN-13 | 978-0-323-29942-8 / 9780323299428 |
| Haben Sie eine Frage zum Produkt? |
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