Automation In Microbiology

Dr Avinash Phadke

With the occurrence of AIDS, there are now proofs of new pathogens causing the disease. This, especially, has necessitated the use of quicker and accurate identification of the bacteria causing these kinds of diseases. Hence, now there is automation of a large number of cultures, especially for blood and sterile body fluids like cerebrospinal fluid.

Most microbiological culture procedures require the use of solid media, like blood agar and MacConkeys agar plates that need to be visually monitored by trained personnel at intervals of 24 hours. These conventional cultures using normal media take at least a minimum of 72 hours to isolate the pathogen and carry out susceptibility test to know the efficacy of antibiotics on simple aerobic bacteria. But this can be completed within 30 hours by using automated techniques. This is especially useful when large number of specimens needs to be cultured, as the instrument, which has been programmed for the same, automatically screens these.

An example of this is the BacT/AlerT 3D culture system. This is the first automated non-radiometric and non-invasive culture system that continuously monitors system for culture of bacteria (both aerobic and anaerobic), fungi and mycobacteria. All these bacteria can be cultured using different media as prescribed. This is a closed system and works on the colorimetric principle of detection of CO2 produced by the organisms. The CO2 causes a lowering of the pH of the medium, which in turn produces a colour change in a sensor attached to the CO2-sensitive base of each bottle. The instrument reacts before this colour change is apparent by means of an audible or visible alert flagged by the computer. The bottles are constantly agitated and are read at 10-minute intervals. The readings are transmitted to a computer compiler, which computes results. This graph can be recalled on a screen any time to monitor production of CO2.

Blood & Body Fluid Cultures

• Blood cultured by the BacT/Alert 3D leads to early detection of pathogens (>89 per cent within 24 hours and 97 per cent within 48 hours) especially in cases of septicaemia, enteric fevers, bacterial endocarditis and other pyrexias of bacterial origin.
• Activated charcoal neutralises antimicrobials and toxins enchancing early recovery of pathogens. Positives are detected faster than Bactec even at low concentrations in blood and body fluids like CSF, CT guided aspirates etc.
• Delayed transport does not compromise results.
• The instrument is capable of recovering significantly more organisms that resin.

Mycobacterial Cultures

• Mycobacterial growth is generally observed within a week in case of smear (1+) positive.
• Speciation into mycobacterium tuberculosis complex and mycobacteria other than tuberculosis takes an additional three days.
• Susceptibility testing to primary line of anti-tuberculosis drugs viz streptomycin, isoniazid, rifampicin, ethambutol and pyrizinamide and secondary line of drugs viz kanamycin, para-amino salicylic acid, cycloserine, ethionamide, capreomycin etc requires 5-10 days.
• Thus, culture would need 10-15 days and susceptibility tests another 5-10 days. Hence, the entire process of culture and susceptibility would take 20-25 days instead of 6-10 weeks taken for conventional culture and susceptibility.
• The system also circumvents problem encountered by the Bactec radiometric system, such as false positives that may occur due to inappropriate threshold values and carryover from positive bottles by inadequately sterilised sample needles.

Automation is particularly useful in the culture of organisms like salmonella, which is a causative agent of typhoid. Conventional cultures will require at least one week monitoring to isolate this organism from blood but by the use of automation the time can be reduced to two to three days.

The percentage of positive cultures is also much higher as the machine is able to recover 88-90 per cent of the disease-producing bacteria as compared to conventional cultures, which is able to recover 35-40 per cent of cultures.

This is useful especially when patients have typhoid and early diagnosis is important, hence both cultures and susceptibility can be done within 30 hours in a case of typhoid. In case the patient is on antibiotics the media has substances like activated charcoal or resins that neutralise the effect of the antibiotic and help to recover the bacterium, which is not possible with conventional methods of culture. Salmonella can be cultured in 90 per cent of cases in the first week, 75 per cent of cases during the second week and 60 per cent during the third week.

For culture of the mycobacterium, which is the causative organism of tuberculosis, automation has proved a boon. This organism requires six to eight weeks to isolate the organism and another six to eight weeks to do a susceptibility test to ascertain the anti- tuberculosis drugs that can kill the organisms. Thus, a culture and susceptibility test would require a minimum of 12-16 weeks. Automation has drastically reduced this period of culture and susceptibility to four to five weeks. Hence, the physician can change the anti-tubercular therapy in case of a patient who has contracted a multi drug resistant strain of tubercle bacilli. So, the time for culture and susceptibility is shortened by a period of at least four to six weeks in case of tuberculosis.

These automated systems also have the capability of increased recovery of organisms. While conventional methods can recover 40-45 per cent of the causative organisms from the specimens the use of automation has increased the recovery of organisms to approx 80-90 per cent from specimens.

The introduction of automation has also helped many microbiologists in ascertaining if the isolate recovered is a pathogen in a particular specimen for example in stool and sputum where normal organisms are also present.

API Systems

• Identification of the isolates will be carried out by integrated systems of routine and automated API systems.
• Susceptibility tests for anaerobes and fungi will be carried out using API systems and against the largest possible range of antibiotics.
• Rapid susceptibility will be carried out for gram negative and staphylococcal isolates on request. These will be reported within 12 hours using API systems. Automation has made it easier to arrive at a precise laboratory diagnosis of infection and helped the physician by providing the antibiogram of the causative organism thus aiding in accurate treatment to the patient.

The writer is Managing Director of  Dr Phadke’s Pathology Lab and Infertility Centre Private Limited, Mumbai.
Email: npilphadkelab@vsnl.net