A 32-year-old man with HIV infection (CD4 count 140 cells/µL) in Imphal has cough, fever and weight loss for 4 weeks. Two sputum samples are negative for acid-fast bacilli on ZN smear, but CBNAAT on one of them reports: MTB detected (low); rifampicin resistance not detected. Which explanation best accounts for the discordant results?

Smear needs about 10,000 bacilli/mL to turn positive

ZN stain does not colour bacilli in HIV patients

CBNAAT also detects non-tuberculous mycobacteria

The CBNAAT cartridge was contaminated by the smear

A 29-year-old woman in Ahmedabad with cough for 5 weeks, previously treated for TB, gives sputum at the designated microscopy centre. CBNAAT reports: MTB detected; rifampicin resistance detected. Mutations in which region does this test detect?

81 base-pair core region of rpoB

Quinolone-resistance region of gyrA

Promoter region of the inhA gene

A 4-month-old girl in a slum in Patna has had bouts of cough for 3 weeks. Each bout ends with an inspiratory whoop or vomiting, and she is well between bouts. Her total leucocyte count is 28,000/µL with 75% lymphocytes. She has received only her birth vaccines. Which specimen and culture medium are best to confirm the diagnosis by culture?

Pernasal swab on charcoal blood agar

Sputum on Lowenstein-Jensen medium

Throat swab on sheep blood agar

Blood sample in brain heart infusion

The operation theatre of a district hospital in Bikaner must sterilise tubes of liquid paraffin and packets of dusting powder used for surgical gloves. Why is the hot air oven preferred over the autoclave for these items?

Steam cannot penetrate oils and powders

Autoclave steam turns paraffin into a toxin

Dry heat kills spores at a lower temperature

Hot air oven cycles are much shorter

Sterilisation, culture media, stains and diagnostics

19 NEET-PG practice questions on Sterilisation, culture media, stains and diagnostics, part of Microbiology. The first 19 are below; the first 5 with answers and explanations (tap Show answer after you have chosen), the rest in Learn with a free account.

Question 1 · Hard
A 45-year-old man in a tribal block of Jharkhand has had cough for 3 weeks and weight loss. At the designated microscopy centre, the medical officer explains his test to the patient and an intern. Consider four statements: 1. CBNAAT detects M. tuberculosis and rifampicin resistance in about two hours. 2. A positive CBNAAT result shows that the bacilli are alive, so it is used to monitor response to treatment. 3. Truenat, an Indian chip-based real-time PCR, is also used under NTEP for diagnosis. 4. Rifampicin resistance on CBNAAT is used as a marker of MDR/RR-TB. Which statements are correct?
1. A1 and 3 only
2. B2, 3 and 4 only
3. C1, 3 and 4 only
4. D1 and 2 only
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Answer: C · 1, 3 and 4 only
Statements 1, 3 and 4 are correct. CBNAAT is a cartridge-based real-time PCR that gives a result for M. tuberculosis and rifampicin resistance in about two hours, and Truenat, a portable chip-based real-time PCR developed in India, is also used under NTEP. Rifampicin resistance is taken as a marker of MDR/RR-TB because it rarely occurs alone, and the specimen then goes for line probe assays and culture-based testing. Statement 2 is false: PCR detects DNA, including DNA from dead bacilli, so it can stay positive during successful treatment; follow-up uses smear microscopy and culture.
- A: Leaves out statement 4; rifampicin resistance is the NTEP marker of MDR/RR-TB.
- B: Includes statement 2; NAAT cannot show whether bacilli are viable.
- D: Includes statement 2; DNA from dead bacilli makes NAAT unsuitable for follow-up.
Question 2 · Hard
A 32-year-old man with HIV infection (CD4 count 140 cells/µL) in Imphal has cough, fever and weight loss for 4 weeks. Two sputum samples are negative for acid-fast bacilli on ZN smear, but CBNAAT on one of them reports: MTB detected (low); rifampicin resistance not detected. Which explanation best accounts for the discordant results?
1. ASmear needs about 10,000 bacilli/mL to turn positive
2. BZN stain does not colour bacilli in HIV patients
3. CCBNAAT also detects non-tuberculous mycobacteria
4. DThe CBNAAT cartridge was contaminated by the smear
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Answer: A · Smear needs about 10,000 bacilli/mL to turn positive
Sputum smear microscopy becomes positive only when there are roughly 5,000-10,000 bacilli per mL, whereas CBNAAT detects a much smaller number of bacilli. People with HIV and low CD4 counts often have paucibacillary, non-cavitary disease, so smear-negative but NAAT-positive TB is common in them. This is why NTEP prefers a NAAT as the first test for people living with HIV. He has bacteriologically confirmed, rifampicin-sensitive TB and should start first-line treatment, with ART timed as per guidelines.
- B: Tubercle bacilli in HIV patients stain normally; they are simply fewer.
- C: CBNAAT targets M. tuberculosis complex DNA and does not detect non-tuberculous mycobacteria.
- D: The two tests are run separately; one does not contaminate the other.
Question 3 · Medium
A 29-year-old woman in Ahmedabad with cough for 5 weeks, previously treated for TB, gives sputum at the designated microscopy centre. CBNAAT reports: MTB detected; rifampicin resistance detected. Mutations in which region does this test detect?
1. A81 base-pair core region of rpoB
2. BQuinolone-resistance region of gyrA
3. CPromoter region of the inhA gene
4. DCodon 315 of the katG gene
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Answer: A · 81 base-pair core region of rpoB
CBNAAT (GeneXpert MTB/RIF) is a cartridge-based real-time PCR that detects M. tuberculosis complex DNA and, with molecular beacons, mutations in the 81-bp rifampicin resistance-determining region of rpoB, the gene for the RNA polymerase beta subunit. Most rifampicin-resistant strains carry a mutation in this region. Because rifampicin resistance usually travels with isoniazid resistance, it is treated as a marker of MDR/RR-TB, and the sample then goes for further drug-susceptibility testing.
- B: gyrA mutations cause fluoroquinolone resistance, read by second-line line probe assay.
- C: inhA promoter mutations cause isoniazid (and ethionamide) resistance, read by line probe assay.
- D: katG codon 315 mutations cause high-level isoniazid resistance, not detected by CBNAAT MTB/RIF.
Question 4 · Medium
A 4-month-old girl in a slum in Patna has had bouts of cough for 3 weeks. Each bout ends with an inspiratory whoop or vomiting, and she is well between bouts. Her total leucocyte count is 28,000/µL with 75% lymphocytes. She has received only her birth vaccines. Which specimen and culture medium are best to confirm the diagnosis by culture?
1. APernasal swab on charcoal blood agar
2. BSputum on Lowenstein-Jensen medium
3. CThroat swab on sheep blood agar
4. DBlood sample in brain heart infusion
Show answerHide answer
Answer: A · Pernasal swab on charcoal blood agar
Paroxysmal cough with whoop, post-tussive vomiting and marked lymphocytosis in an unimmunised infant point to pertussis. Bordetella pertussis attaches to ciliated cells of the nasopharynx, so a pernasal (nasopharyngeal) swab is taken and plated on Regan-Lowe charcoal blood agar or Bordet-Gengou medium, which neutralise inhibitory fatty acids. Culture is most often positive in the catarrhal and early paroxysmal stages, and PCR on the same swab is more sensitive.
- B: Lowenstein-Jensen medium is for mycobacteria, and infants cannot produce sputum.
- C: A throat swab misses the organism, and plain blood agar does not support its growth.
- D: Pertussis is not a bacteraemic infection, so blood culture is not useful.
Question 5 · Medium
The operation theatre of a district hospital in Bikaner must sterilise tubes of liquid paraffin and packets of dusting powder used for surgical gloves. Why is the hot air oven preferred over the autoclave for these items?
1. AAutoclave steam turns paraffin into a toxin
2. BSteam cannot penetrate oils and powders
3. CDry heat kills spores at a lower temperature
4. DHot air oven cycles are much shorter
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Answer: B · Steam cannot penetrate oils and powders
Moist heat works only where steam can reach and condense, and anhydrous materials such as oils, greases and powders repel water. Dry heat in a hot air oven, usually 160 °C for 2 hours, penetrates them and kills microbes by oxidation and protein denaturation. The oven is also used for glassware and metal instruments, but not for rubber or plastics, which melt.
- A: Steam does not make paraffin toxic; it simply fails to penetrate it.
- C: Dry heat needs a higher temperature and longer time than moist heat to kill spores.
- D: The hot air oven cycle is longer, not shorter, than the autoclave cycle.
Question 6 · Medium
The laboratory of a medical college in Siliguri receives CSF from an 18-month-old girl with bacterial meningitis whose family had refused vaccines. Pleomorphic gram-negative coccobacilli are seen, and on sheep blood agar pinpoint colonies appear only close to a streak of Staphylococcus aureus. What does the staphylococcus supply to the organism?
1. ABoth X and V factors
2. BCysteine and iron
3. CV factor (NAD)
4. DX factor (haemin)
Answer and explanation in LearnFree account
Question 7 · Easy
At a leprosy clinic in Raipur, a slit-skin smear is taken from the ear lobe of a 38-year-old man with infiltrated, shiny facial skin and thickened ulnar nerves. The smear is stained by the Ziehl-Neelsen method. What decolouriser should be used for this smear?
1. A5% sulphuric acid
2. B1% sulphuric acid
3. CAcetone-alcohol
4. D20% sulphuric acid
Answer and explanation in LearnFree account
Question 8 · Medium
The TB culture laboratory of a medical college in Bhopal prepares its own Lowenstein-Jensen slopes for samples from patients with presumptive drug-resistant TB. The medium contains egg, which must solidify without being destroyed. How is this medium sterilised?
1. AAutoclaving at 121 °C for 15 minutes
2. BTyndallisation by steaming at 100 °C
3. CInspissation at 80-85 °C on three days
4. DMembrane filtration through 0.22 µm
Answer and explanation in LearnFree account
Question 9 · Medium
A microbiology intern in Mysuru stains a CSF deposit from a febrile infant and reviews the Gram method with her tutor. Consider four statements: 1. Crystal violet is the primary stain. 2. Gram's iodine acts as a mordant that fixes the dye in the cell. 3. Gram-negative cells lose the dye because their thin peptidoglycan and outer membrane lipid let the decolouriser extract it. 4. Safranin, the counterstain, gives gram-positive cells their red colour. Which statements are correct?
1. A1, 2 and 4 only
2. B1, 2 and 3 only
3. C1 and 3 only
4. D1 and 2 only
Answer and explanation in LearnFree account
Question 10 · Hard
After three hostel students in Jaipur develop culture-proven enteric fever, the district surveillance team screens the hostel cooks. One cook had typhoid two years ago and feels well. A stool sample is collected from him. Into which broth should it be placed for enrichment before plating?
1. AAlkaline peptone water
2. BTaurocholate bile broth
3. CThioglycollate broth
4. DSelenite F broth
Answer and explanation in LearnFree account
Question 11 · Hard
A 40-year-old man in Kozhikode with chronic otitis media has fever, headache and a ring-enhancing lesion in the temporal lobe. Pus aspirated at surgery is foul-smelling. Its Gram stain shows many organisms of mixed shapes, but aerobic culture on blood and MacConkey agar is sterile after 48 hours. How should the laboratory culture this specimen?
1. ASabouraud dextrose agar with chloramphenicol
2. BRobertson's cooked meat medium with anaerobic incubation
3. CLowenstein-Jensen medium incubated for eight weeks
4. DBlood agar incubated aerobically for seven days
Answer and explanation in LearnFree account
Question 12 · Easy
During monsoon floods in a relief camp near Kolkata, a 30-year-old woman passes profuse rice-water stools and is severely dehydrated. A stool sample is sent to the district laboratory. On which selective plating medium should it be inoculated to isolate the likely organism?
1. ASkirrow campylobacter selective agar
2. BThiosulphate citrate bile salt sucrose agar
3. CModified Thayer-Martin agar
4. DWilson and Blair bismuth sulphite agar
Answer and explanation in LearnFree account
Question 13 · Medium
A 58-year-old man is admitted to a Lucknow hospital with community-acquired pneumonia. Two sets of blood cultures were drawn from two separate sites before antibiotics. After 3 days, one bottle of one set grows coagulase-negative staphylococci; the other three bottles remain sterile. He has no prosthetic device or central line. How should this result be interpreted?
1. ATrue bacteraemia needing vancomycin
2. BInfective endocarditis until proved otherwise
3. CLaboratory error needing a repeat Gram stain
4. DProbable skin contaminant during collection
Answer and explanation in LearnFree account
Question 14 · Easy
Which holding time and temperature make up the standard autoclave cycle for culture media and linen at 15 lb per square inch?
1. A160 °C for 2 hours
2. B134 °C for 30 minutes
3. C100 °C for 20 minutes
4. D121 °C for 15 minutes
Answer and explanation in LearnFree account
Question 15 · Easy
The central sterile supply department of a district hospital in Nashik validates its steam autoclave every week with a biological indicator strip placed in the centre of the load. Spores of which organism should the strip contain?
1. AGeobacillus stearothermophilus
2. BBacillus cereus
3. CClostridium tetani
4. DBacillus atrophaeus
Answer and explanation in LearnFree account
Question 16 · Medium
A 47-year-old renal transplant recipient in Chennai on tacrolimus and steroids has a cavitating lung lesion and a ring-enhancing brain lesion. Gram stain of bronchoalveolar lavage fluid shows thin, beaded, branching gram-positive filaments. Which staining method best separates the likely organism from Actinomyces?
1. AIndia ink preparation
2. BAlbert stain for volutin granules
3. CModified ZN stain with 1% sulphuric acid
4. DStandard ZN stain with 20% sulphuric acid
Answer and explanation in LearnFree account
Question 17 · Medium
A 26-year-old truck driver attends the STI clinic of a medical college in Nagpur with dysuria and a thick yellow urethral discharge for 3 days. Gram stain of the discharge shows many pus cells containing gram-negative kidney-shaped diplococci. On which medium should the urethral swab be cultured to isolate the organism?
1. AMacConkey agar
2. BLoeffler serum slope
3. CModified Thayer-Martin agar
4. DBordet-Gengou agar
Answer and explanation in LearnFree account
Question 18 · Medium
A 34-year-old woman in Pune has fever and myalgia during an influenza season. A nasopharyngeal swab is sent for real-time RT-PCR for influenza A (H1N1). Which step must take place before the PCR can amplify the viral target?
1. AReverse transcription of RNA into cDNA
2. BLigation of viral DNA fragments
3. CCutting of viral DNA by restriction enzymes
4. DTranscription of viral DNA into mRNA
Answer and explanation in LearnFree account
Question 19 · Medium
A flexible bronchoscope in a district hospital in Thrissur is used for several patients on the same morning, including one later found to have sputum-positive tuberculosis. The scope cannot withstand heat. After thorough manual cleaning, what is the most appropriate way to reprocess it before the next patient?
1. AAutoclaving at 121 °C
2. BImmersion in 2% glutaraldehyde
3. CWiping with 70% ethyl alcohol
4. DHot air oven at 160 °C
Answer and explanation in LearnFree account

Sterilisation, culture media, stains and diagnostics

A 29-year-old woman in Ahmedabad with cough for 5 weeks, previously treated for TB, gives sputum at the designated microscopy centre. CBNAAT reports: MTB detected; rifampicin resistance detected. Mutations in which region does this test detect?

The operation theatre of a district hospital in Bikaner must sterilise tubes of liquid paraffin and packets of dusting powder used for surgical gloves. Why is the hot air oven preferred over the autoclave for these items?

At a leprosy clinic in Raipur, a slit-skin smear is taken from the ear lobe of a 38-year-old man with infiltrated, shiny facial skin and thickened ulnar nerves. The smear is stained by the Ziehl-Neelsen method. What decolouriser should be used for this smear?

The TB culture laboratory of a medical college in Bhopal prepares its own Lowenstein-Jensen slopes for samples from patients with presumptive drug-resistant TB. The medium contains egg, which must solidify without being destroyed. How is this medium sterilised?

After three hostel students in Jaipur develop culture-proven enteric fever, the district surveillance team screens the hostel cooks. One cook had typhoid two years ago and feels well. A stool sample is collected from him. Into which broth should it be placed for enrichment before plating?

During monsoon floods in a relief camp near Kolkata, a 30-year-old woman passes profuse rice-water stools and is severely dehydrated. A stool sample is sent to the district laboratory. On which selective plating medium should it be inoculated to isolate the likely organism?

Which holding time and temperature make up the standard autoclave cycle for culture media and linen at 15 lb per square inch?

The central sterile supply department of a district hospital in Nashik validates its steam autoclave every week with a biological indicator strip placed in the centre of the load. Spores of which organism should the strip contain?

A 34-year-old woman in Pune has fever and myalgia during an influenza season. A nasopharyngeal swab is sent for real-time RT-PCR for influenza A (H1N1). Which step must take place before the PCR can amplify the viral target?