The review searched PubMed/MEDLINE and Europe PMC from database inception through 18 August 2026, supplemented by citation chaining and targeted retrieval of guidance and comparator studies. It evaluated complete specimen-to-result systems rather than isolated nuclease reactions, classifying studies by evidence stage and appraising them across direct intended-use evidence, specimen integration, measurement validity, control and containment, and operational or clinical utility. Because the literature varied substantially in analytes, specimen matrices, amplification methods, outputs and study designs, findings were synthesized narratively rather than pooled. The review was not prospectively registered as a systematic review, retrospective screening counts were not reconstructed, and relevant engineering papers outside the selected bibliographic routes may have been missed.

Existing antenatal pathways already rely on established serological and antigen-based screening for HIV, HBV and syphilis. Molecular testing becomes most defensible when it resolves a specific downstream decision that a screen cannot: calibrated HIV viral-load assessment, early infant diagnosis, HBV risk classification after a positive hepatitis B surface antigen result, or direct detection of Treponema pallidum in selected early or discordant presentations. The timing of transmission and the interventions differ across the three infections, so their diagnostic requirements cannot be treated as interchangeable. HIV may be transmitted during pregnancy, labor, delivery or breastfeeding; HBV transmission is strongly associated with maternal viremia around delivery; and T. pallidum can cross the placenta, making timely maternal treatment central to preventing congenital outcomes.

HIV EID had the most compelling development case because maternal antibodies make early serology unsuitable, infant prophylaxis can reduce viral abundance, and treatment should begin promptly when infection is diagnosed. A deployable assay would need to recover low-copy HIV nucleic acid from metered microvolume whole blood, dried blood spots or a separately validated plasma workflow, while remaining inclusive across relevant HIV-1 subtypes and recombinants. The most directly relevant study identified in the review used an RPA–Cas12a assay targeting the HIV-1 clade C pol region. Among 28 infant plasma specimens from children younger than two years, 27 were positive and one inconclusive, yielding a positive-result proportion of 96.4% when the inconclusive result remained in the denominator; the overall study reported 96% sensitivity and 92.65% specificity. The lowest detected clinical sample was estimated at approximately 3,000 copies/mL, but the workflow required plasma RNA extraction and reverse transcription and did not test dried blood spots or microvolume whole blood.

The HIV results demonstrate detection feasibility rather than readiness to replace validated qualitative nucleic acid testing. Infant studies need to report performance in the lowest reference viral-load strata, effects of prophylaxis, initial invalid and repeat rates, specimen adequacy, and the time from testing to treatment. Maternal viral-load monitoring is less aligned with current CRISPR evidence because management may require longitudinal assessment of suppression, rebound and change, with results interpreted in copies per millilitre. Digital proof-of-concept systems have illustrated alternative approaches, but small evaluated volumes, subsampling noise, signal saturation and uncertain calibration constrain them. In one membrane-based digital Cas13a study, 20 patient plasma specimens produced 60% sensitivity and 100% specificity, with an estimated end-to-end detection limit of about 2,000 copies/mL, despite close tracking of expected input in spiked material. The investigators therefore positioned maternal HIV CRISPR testing as an investigational triage tool at most, rather than a replacement for calibrated viral-load measurement.

Maternal HBV reflex testing had the largest body of clinical-specimen evidence, yet it also exposed the difference between detecting viral DNA and making a prophylaxis decision. Under guidance cited by the review, tenofovir disoproxil fumarate prophylaxis applies for HBsAg-positive pregnant women with HBV DNA at or above 200,000 IU/mL or positive HBeAg, while a broader pathway may be considered when those markers are unavailable. A CRISPR classifier would therefore need to report below-threshold, gray-zone, above-threshold and out-of-range results calibrated to the internationally traceable unit, rather than merely report that HBV DNA was detected. Validation cohorts would need to deliberately include specimens near the applicable decision boundary and assess genotype, operator, reagent-lot, high-load and matrix effects. A one-pot LAMP–Cas12b study detected 198 of 200 qPCR-positive clinical specimens and all 36 qPCR-negative specimens, yet detection was 16 of 16 for genotype C and seven of nine for genotype B, with guide-region mutations found in both false negatives. This signal of inclusivity failure shows why aggregate accuracy cannot substitute for boundary-focused validation.

Other HBV systems have advanced one-pot operation, whole-blood processing and digital measurement, but none established the required pregnancy use case. A multistage PCR–T7–Cas13a assay reported positive agreement of 100% in 90 qPCR-positive serum specimens and negative agreement of 94.7% in 190 qPCR-negative specimens, while a low-level-viremia cohort showed that detection became less reliable below 100 IU/mL. A digital Cas12a platform produced concentration-responsive results but slightly lower estimates than droplet digital PCR and did not establish interchangeability through Bland–Altman agreement analysis. Maternal classification also depends on the clinical consequence of error: missing an eligibility threshold may deny prophylaxis, whereas an erroneous positive may trigger unnecessary treatment. The evidence therefore supports rapid reflex access as a development objective, but not a claim that CRISPR HBV testing is ready for pregnancy care.

For syphilis, the review found the narrowest defensible role for CRISPR direct detection. Antenatal prevention remains centered on serological screening followed by adequate penicillin treatment, which is generally available during the same visit. Direct pathogen testing is most plausible as an adjunct when a compatible lesion is present during a possible seronegative window, when serology is discordant, or when reinfection is suspected and an appropriate lesion or mucosal specimen can be collected. A PCR–T7–LwCas13a clinical benchmark detected T. pallidum in 93.3% of a mixed specimen set with 100% specificity, but sensitivity in whole blood was 71.4% in primary syphilis and 87.7% in secondary syphilis, substantially below the yield from skin biopsies and genital lesion swabs. These mixed estimates cannot be extrapolated to pregnancy, latent infection or congenital disease. A negative direct test may reflect sparse or intermittently sampled organisms, while a positive result after treatment may reflect residual nucleic acid rather than viable infection, treatment failure or transmissibility.

Complete-system design remains central to translation. Cas12- and Cas13-based assays typically pair sequence-specific recognition with PCR, recombinase polymerase amplification, recombinase-aided amplification, LAMP or another amplification step and a fluorescent, colorimetric or lateral-flow readout. Each additional transfer can increase carryover risk and complicate quality control, while a portable strip attached to an open amplification tube is not a sample-to-answer device. Decentralized assays need explicit controls for specimen adequacy, extraction and amplification performance, and CRISPR or reader function, together with contamination controls and predefined invalid-result rules. Readouts should distinguish positive, negative, invalid and indeterminate outcomes, and quantitative assays need reportable ranges, calibration and measurement uncertainty. The review also warned that one-pot operation does not necessarily mean simultaneous chemistry, closed containment or full automation. For HIV EID, a shared service platform with a validated qualitative assay appears more defensible than a triplex that compromises recovery or control.

Overall, the review concluded that clinical priority and evidence maturity diverge. HIV EID is the most compelling near-term opportunity, provided prospective studies validate microvolume or dried-blood-spot workflows, prophylaxis-exposed infants, viral diversity, integrated controls and time to treatment. Maternal HBV has the strongest clinical-specimen base but needs IU/mL-traceable, boundary-focused pregnancy validation, while syphilis direct detection should remain a specimen-specific adjunct with explicit limits on excluding infection. Future development should proceed through analytical characterization of a locked configuration, prospective evaluation in the intended population, multisite testing by intended users, and pragmatic assessment of whether results improve care equitably. Until those stages are completed, the identified CRISPR systems are better regarded as developmental components and narrowly targeted candidates than as replacements for established HIV, HBV and syphilis pathways.

Subject of Research: Clinical roles and translational gaps of CRISPR diagnostics for preventing mother-to-child HIV, hepatitis B and syphilis transmission

Article Title: CRISPR diagnostics for maternal–infant HIV, hepatitis B, and syphilis care: clinical roles and translational gaps

Article References: Luo, H., Lin, L., Luo, F., Huang, X., Yu, Q., Li, Q., & Zha, H. (2026). CRISPR diagnostics for maternal–infant HIV, hepatitis B, and syphilis care: clinical roles and translational gaps. Frontiers in Microbiology, 17, Article 1977927. https://doi.org/10.3389/fmicb.2026.1977927

Image Credits: AI Generated

DOI: 10.3389/fmicb.2026.1977927

Keywords: CRISPR diagnostics, HIV early infant diagnosis, hepatitis B virus, syphilis, mother-to-child transmission, molecular testing, clinical validation, viral diagnostics, CRISPR, diagnostics, maternal, infant