Stephen H. Gillespie - Medical Microbiology and Infection at a Glance

Pathogen identification is important because it can predict disease and prognosis, e.g. Vibrio cholerae causes severe watery diarrhea and is potentially fatal.

Identification of some organisms requires prompt public health action: e.g. contact tracing for patients with meningococcal disease.

Bacterial identification depends on colonial morphology on agar, microscopic morphology, and biochemical tests. Matrix‐assisted‐laser‐desorption/ionization‐time‐of‐flight (MALDI‐TOF) can achieve this in 20 minutes. Nucleic acid amplification tests (NAATs) and gene sequencing are used especially when organisms are slow growing (e.g. Mycobacterium tuberculosis ) or impossible to grow (e.g. Trophyrema whippelii ).

Susceptibility testing (DST) determines whether treatment is likely to be successful remembering that clinical response depends on host factors too. A susceptible organism should respond to a standard dose of an antimicrobial; an intermediate resistant strain should respond to a larger dose; and a resistant organism is likely to fail therapy with that antibiotic.

DST can be achieved by measuring the diameter of an inhibition zone around a paper disc with incorporated antibiotics. Susceptibility is defined by a ‘breakpoint’ in growth. These methods are standardized by international bodies such EUCAST to ensure reproducibility. Automated methods can achieve this more rapidly.

The minimum inhibitory concentration, which is the lowest dose that completely inhibits growth, is a more objective method and enables resistance levels to be related to the concentration of antibiotic that is achievable in the tissues.

Susceptibility can be assessed rapidly by hybridization or sequence‐based methods that detect specific antibiotic‐resistance mutations.

An infection can be diagnosed by detecting the immune response to the pathogen: for example by detection of rising or falling antibody concentrations more than a week apart, or by the presence of a specific IgM or specific pathogen antigen. Detecting the activity of specific T‐cells can provide evidence of exposure to tuberculosis. Serological techniques are used for organisms that are difficult or impossible to grow such as viruses (e.g. HIV or hepatitis B).

A labelled DNA probe can bind to components of the pathogen and be detected by the activity of its attached label. This technique is specific and rapid, but less sensitive because there are no amplification steps.

Nucleic acid amplification tests (NAATs) amplify specific regions of the pathogen genome (DNA or RNA) to make the diagnosis until there is sufficient for detection. Primers are designed to bind to target a specific pathogen sequence and a polymerase synthesizes new nucleic acid over multiple cycles. Automated systems and commercial kits can follow this process in real time and make these tests available in many laboratories. NAATs have the advantage that they can detect slow or difficult to grow organisms, or make a diagnosis in the patient who has taken antibiotics. PCR can also detect virulence determinants or resistance determinants creating a surrogate susceptibility result (e.g. rpo B gene mutation for rifampicin resistance in M. tuberculosis ).

The reducing cost of sequencing the whole pathogen sequence means that this is increasingly widely available. It provides extremely detailed information on the pathogen. Using complex manipulations of the genomic data it is possible to determine the relationship between organisms and identify their transmission. It has become vital in tracking viruses like influenza, and resistant or virulent bacteria in community or hospital outbreaks. Rapid availability of data can rapidly rule in or rule out an outbreak.

Antibiotics aim to kill organisms while causing no harm to the patient – this concept is known as selective toxicity. It is best achieved by inhibiting a pathogen function that human cells do not have. Bacteria have a rigid cell wall with peptidoglycan that human cells lack and can be inhibited by penicillin. Finding selective targets is more difficult for eukaryotic pathogens that can have similar pathways to humans. Similarly challenging are obligate intracellular pathogens such as viruses where detailed knowledge of viral reproduction is required to create effective treatments.

The difference between the dose necessary for treatment and that which causes harm is usually large and is known as the therapeutic index. The aminoglycosides are exceptions to this because doses just above the therapeutic level can be toxic. While all antimicrobials have potential unwanted effects, fortunately serious unwanted effects are not frequent. We aim, therefore, to provide an effective antibiotic regimen that maximizes the chance of a successful outcome with the least risk of adverse events.

Mild gastrointestinal upset is probably the most frequent side effect of antibiotic therapy. Antibiotic activity can upset the balance of the normal flora within the gut (β‐lactams are especially likely to do this) resulting in overgrowth of commensal organisms such as Candida spp. Alternatively, antibiotic therapy may provoke diarrhoea or, more seriously, pseudomembranous colitis (see Chapter 24).

Cutaneous manifestations range from mild urticaria or maculopapular, erythematous eruptions to erythema multiforme and the life‐threatening Stevens–Johnson syndrome. Most cutaneous reactions are mild and resolve after discontinuation of therapy.

Patients receiving chloramphenicol or antifolate antibiotics may exhibit dose‐dependent bone marrow suppression. More seriously, aplastic anaemia may rarely complicate chloramphenicol therapy. High doses of β‐lactam antibiotics may induce granulocytopenia. Antibiotics are a rare cause of haemolytic anaemia. Many antibiotics cause a mild reversible thrombocytopenia or bone marrow depression.

Aminoglycosides may cause renal toxicity by damaging the cells of the proximal convoluted tubule. Patients who are elderly, have pre‐existing renal disease or are also receiving other drugs with renal toxicity are at higher risk. Tetracyclines may also be toxic to the kidneys.

Isoniazid and rifampicin may cause hepatitis; this is more common in patients with pre‐existing liver disease. Other agents associated with hepatitis are tetracycline, erythromycin, pyrazinamide, ethionamide and, very rarely, ampicillin or fluoroquinolones. Cholestatic jaundice may follow tetracycline or high‐dose fusidic acid therapy.

Allergy to antibiotics is relatively common with a wide spectrum of impacts. Anaphylaxis from beta‐lactam antibiotics is potentially life‐threatening, but rare. A history of penicillin anaphylaxis must be captured on a patient’s medical record. Severe skin and systemic reactions such as Stevens–Johnson syndrome associated, for example, with co‐trimoxazole, can also be life‐threatening. Other allergic reactions can include skin reactions such as fixed drug eruptions.