Semen Analysis According to the WHO 6th Edition: What Changed and Why It Matters

Semen analysis is the cornerstone of male fertility evaluation, and its value depends on doing it in a standardised way. In 2021 the World Health Organization published the 6th edition of its Laboratory Manual for the Examination and Processing of Human Semen, replacing the 5th edition from 2010. Here is what changed and what it means for andrology laboratories.

Key changes in the 6th edition

1. Reference values reframed

The 6th edition presents reference distributions drawn from a larger and more geographically diverse population of men whose partners conceived within 12 months. Importantly, the manual emphasises that the lower reference limits (the 5th centiles) are not strict cut-offs between fertile and infertile men. Semen parameters should be interpreted in the full clinical context of the couple.

2. Motility categories restored

The 5th edition grouped motility into progressive, non-progressive and immotile. The 6th edition reintroduced the distinction between rapidly progressive and slowly progressive motility, as rapid progressive motility carries useful clinical information. Accurate assessment requires a properly controlled temperature.

3. Clear separation of basic, extended and advanced tests

The manual organises tests into:

• Basic examination - the standard semen analysis
• Extended examination - such as tests for antisperm antibodies or markers of inflammation, when clinically indicated
• Advanced examination - research-oriented or specialised tests, such as sperm DNA fragmentation, where the evidence and standardisation are still developing

4. Stronger emphasis on quality

The manual places greater emphasis on standardised methods, internal and external quality control, and staff training - recognising that variation between laboratories has long limited the usefulness of semen analysis.

Lower reference limits (5th centile) in the 6th edition

These are the commonly cited lower reference limits from the 6th edition:

• Semen volume: 1.4 mL
• Sperm concentration: 16 million per mL
• Total sperm number: 39 million per ejaculate
• Total motility (progressive + non-progressive): 42%
• Progressive motility: 30%
• Vitality: 54% live
• Normal forms (morphology): 4%

Remember: a value below these limits does not by itself mean a man is infertile, and values above them do not guarantee fertility.

Practical implications for laboratories

• Collection: clear patient instructions on abstinence period and complete collection; record the time of collection and any spillage
• Liquefaction and timing: begin analysis within the recommended window after collection
• Counting: use the recommended dilution and counting chambers; count enough sperm for statistical reliability
• Motility: assess at a controlled temperature and classify into rapid, slow, non-progressive and immotile
• Morphology: use strict criteria with proper staining, and train staff with regular comparisons
• Reporting: report results clearly with the method used, so clinicians can interpret them correctly
• Quality control: internal QC and participation in external quality assessment schemes

Why standardisation matters

When semen analysis is done inconsistently, couples may be sent for unnecessary treatment - or miss treatment they need. Standardised, quality-controlled testing protects patients and gives clinicians results they can trust.

Train in WHO 6th edition protocols

ReproNest's Advanced Andrology & Semen Analysis Certification (WHO 6th Edition Protocols) provides hands-on training in every step of semen analysis, sperm preparation and quality control. Read about building a career in andrology, or contact our team.