Beyond the homozygous paradigm: symptomatic partial biotinidase deficiency in a heterozygous child—first case report from Nepal

Abstract

Background

Biotinidase deficiency (BTD) is a rare genetic condition inherited in an autosomal recessive pattern that affects multiple systems. Biotinidase (EC 3.5.1.12) cleaves the vitamin, biotin, from the biocytin and the dietary protein-bound sources, and recycles the biotin. It manifests with a range of neurocutaneous symptoms, including seizures, hypotonia, ataxia, skin rashes, alopecia, hearing loss, optic atrophy, and metabolic crises that resemble sepsis, delay in identification and prompt treatment, results in irreversible brain damage, coma, and, in severe cases, death. Newborn screening can help in early diagnosis as it is amenable to treatment with pharmacological doses of biotin. There has been no study from Nepal about Biotinidase deficiency highlighting the clinical features, diagnosis and response to treatment.

Case presentation

An 18-month-old male, born at term via normal delivery to non-consanguineous parents, with a normal neonatal period, presented to Kanti Children’s Hospital (KCH) with a 3-month history of multiple abnormal body movements and developmental regression. There were no known familial or genetic illnesses. Developmentally, he achieved milestones appropriately until 12 months of age, since then 15 months of age he had development regression. Ophthalmology study showed bilateral pallor of optic disc. MRI, EEG was done thinking of the inborn error of metabolism. A whole exome sequencing identified a heterozygous pathogenic variant in the BTD gene (c.38_44delinsTCC, p.Cys13Phefs*36), and retrospective enzyme assay confirmed partial biotinidase deficiency (3.20 nmol/min/mL; ~25% of mean normal activity). Prompt clinical improvement following biotin supplementation supported the diagnosis despite a non-specific initial presentation. He was then managed with oral biotin 10 mg and supportive care. Follow-up assessments at 3, 6, and 12 months post-treatment showed remarkable improvement.

Conclusions

This case highlights the clinical and diagnostic challenges of biotinidase deficiency (BTD) in resource-limited settings like Nepal, where the absence of newborn screening and limited molecular diagnostic capacity delay timely intervention. Increased clinician awareness, and local research to better understand the spectrum of BTD mutations, their clinical implications and to analyze the effectiveness of implementation of newborn screening programs. Early recognition and treatment of BTD can significantly improve patient outcomes and reduce the burden of this potentially devastating yet treatable disorder.

Clinical trial number

Not applicable.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12887-025-05822-2.

Background

Biotinidase deficiency (BTD) is a rare autosomal recessive metabolic disorder that affects multiple organ systems [1]. Its global incidence ranges from 1 in 40,000 to 1 in 60,000 live births [2]. Biotinidase (EC 3.5.1.12) is an enzyme responsible for cleaving biotin from biocytin and dietary protein-bound sources, thereby recycling biotin for reuse [1]. The BTD gene is located on chromosome 3p25 and comprises four exons spanning a total of 1,629 base pairs [3]. More than 200 mutations have been identified in the BTD gene to date [4].

Diagnosis is primarily based on measuring biotinidase enzyme activity in plasma or serum [1]. Elevated levels of urinary organic acids such as 3-hydroxyisovaleric acid, 3-hydroxypropionic acid, lactic acid, and 3-methylcrotonyl glycine in untreated patients can support the diagnosis [5]. Whole genome sequencing (WGS) can also confirm the diagnosis by identifying pathogenic variants in the BTD gene.

BTD can present at any age, from infancy to adulthood, although it is more commonly identified in infancy [6]. The clinical spectrum includes neurocutaneous symptoms such as seizures, hypotonia, ataxia, skin rashes, alopecia, hearing loss, optic atrophy, and metabolic crises that may mimic sepsis. Delayed diagnosis and treatment can lead to irreversible brain damage, coma, and even death in severe cases [7]. Typically, clinical manifestations occur in individuals with homozygous or compound heterozygous mutations, whereas heterozygous carriers are usually asymptomatic [1]. However, in this report, we describe a symptomatic patient with heterozygous biotinidase deficiency, an unusual presentation.

Newborn screening plays a crucial role in early diagnosis, as the condition is treatable with pharmacological doses of biotin [2, 6]. However, no studies from Nepal have yet documented the clinical features, diagnostic approaches, or treatment outcomes of patients with BTD [8].

Here, we present the first reported case of biotinidase deficiency from Nepal, highlighting the clinical presentation, diagnostic process, and response to treatment.

Case presentation

Clinical presentation

An 18-month-old male, born at term via normal vaginal delivery to non-consanguineous parents, with an unremarkable neonatal period, presented to Kanti Children’s Hospital with a three-month history of multiple abnormal body movements and developmental regression. The child was exclusively breastfed until six months of age, followed by the gradual introduction of complementary feeding. There was no known family history (Supplementary file S1) of genetic or neurological disorders. Developmental milestones were appropriate until 12 months of age, at which point he was able to stand with support, walk with occasional falls, and attempt independent standing. He demonstrated a mature pincer grasp, responded to his name, engaged in simple ball games, vocalized 1–2 meaningful words, recognized sounds, and oriented toward their source, reflecting a Development Quotient (DQ) of approximately 100, consistent with normal developmental progress at that age.

The first seizure episode occurred at seven months of age and was associated with fever. By 13 months, he had experienced three episodes of generalized tonic-clonic seizures (GTCS). At 12 months, he had an episode of status epilepticus requiring intervention at another center. At 15 months, he was admitted to the pediatric intensive care unit (PICU) at our center due to a lower respiratory tract infection and GTCS. Following this admission, he exhibited significant developmental regression, losing previously acquired motor, cognitive, and social milestones. He was unable to walk independently, creep upstairs, initiate scribbling, speak words, or imitate parental activities. His mother reported that he only adjusted his head position to follow objects instead of actively tracking them. This regression corresponds to a DQ of 50–55, indicating moderate to severe developmental delay.

At the presentation, the child exhibited spontaneous limb movements and cooing but lacked neck holding and rolling over. There were no noted comorbidities. Anthropometric measurements revealed a weight of 9 kg (10th percentile), length of 76 cm (15th percentile), and head circumference of 45 cm (10th percentile).

Neurological examination showed that the child was awake but unresponsive to his surroundings. Examination of the skull, neck, and spine revealed no signs of meningeal irritation, malformations, injuries, pain, or tenderness. Cranial nerve assessment showed normal pupillary response to light and the ability to follow objects. Fundoscopic examination revealed bilateral optic disc pallor. Facial symmetry was preserved with an intact smile response. The child responded to auditory stimuli and was able to swallow milk. The tongue appeared normal without fasciculations or asymmetry.

Motor examination revealed symmetrical muscle bulk without fasciculations or involuntary movements. Muscle tone was decreased (hypotonia) in all flexor and extensor groups. Muscle power was 2/5 in both the upper and lower extremities. Bilateral deep tendon reflexes at the knee and ankle were hyperreflexic. Clonus was absent, and the bilateral plantar response was mute. Systemic examination revealed no abnormalities.

Investigations

The working diagnosis of neurodegenerative delay prompted an extensive evaluation. EEG revealed an abnormal sleep pattern with epileptiform discharges in the right centro-temporal region, suggestive of encephalopathy. Brain MRI Axial T2 section at level of corpus callosum (Figure 1) demonstrated prominent frontal sulcal spaces, consistent with benign enlargement of the subarachnoid spaces in infancy (BESSI), along with delayed myelination of the rostrum, genu, and trunk/body of the corpus callosum. Ophthalmological examination revealed bilateral optic disc pallor, while hearing assessment was normal. Laboratory tests, including thyroid function, vitamin D levels, and electrolytes, were within normal limits. Arterial blood gas (ABG) analysis performed during the ICU stay showed a pH of 7.30 (normal range: 7.35–7.45), indicating metabolic acidosis with elevated lactate levels.

Fig. 1.

Axial T2 section at level of corpus callosum of brain showing: Widening of bifrontal (red arrow) and anterior inter hemispheric CSF spaces (black arrow) without flattening of adjacent Gyri, Normal sulci posteriorly and normal ventricular size with no pressure effects on surrounding brain tissue. Normal cortical veins within. Normal white matter myelination in splenium (yellow arrow) and non-myelination of body (*) and genu (x) of corpus callosum

Whole Exome Sequencing (WES) was performed on an EDTA blood sample using next-generation sequencing technology. Library preparation and sequencing were conducted on an Illumina platform, achieving an average read depth of > 30×. Reads were aligned to the human reference genome GRCh38/hg38, utilizing the alt-masked version to enhance variant calling accuracy in complex genomic regions. Variant calling was executed using the Genome Analysis Toolkit (GATK) Best Practices pipeline. Subsequent variant annotation incorporated data from ClinVar, gnomAD, and the Human Gene Mutation Database (HGMD). For the BTD gene, transcript ID NM_001370658.1 was employed. Variant classification was adhered to the American College of Medical Genetics and Genomics (ACMG) guidelines.

Whole-exome sequencing (WES) revealed a heterozygous pathogenic variant in the biotinidase (BTD) gene (c.38_44delinsTCC, p.Cys13Phefs*36) (Table 1) (Supplementary File S1). Although individuals with heterozygous biotinidase deficiency typically remain asymptomatic, the reversible and treatable nature of this condition warranted enzyme activity testing while conducting workup for other differential diagnoses. Biotinidase activity was retrospectively measured and found to be 3.20 nmol/min/mL (normal > 5.00 nmol/min/mL; mean normal activity 7.2 ± 1.1 nmol/min/mL). According to biochemical classification, enzyme activity below 10% of the mean normal indicates profound deficiency, while levels between 10 and 30% are consistent with partial deficiency. The measured value corresponded to approximately 25% of mean normal activity, confirming a partial biotinidase deficiency. Notably, there was marked clinical improvement soon after initiating biotin supplementation, supporting a definitive diagnosis of biotinidase deficiency despite the initially non-specific presentation.

Table 1.

Whole exome sequencing using EDTA blood sample

Gene	Disease and Inheritance (OMIM phenotype number)	Genomic Position	Variant details
BTD	Biotinidase deficiency (AR - Autosomal Recessive) (OMIM ID- 253260)	Chr3:g.15635477_15635483	Exon/ Intron No: Exon 2
			Nucleotide change: c.38_44delinsTCC
			Amino Acid Change: (p.Cys13Phefs*36)
			Transcript Id: NM_001370658.1
Molecular consequence	Allele Frequency	Zygosity	ACMG Classification
Frameshift	1000 Genome: Novel	Heterozygous	Pathogenic (PVS1, PM2, PP5)
	gnomAD: 0.055%

Management and outcome

The child was started on oral biotin at a dose of 10 mg daily, based on clinical guidelines for biotinidase deficiency. No other pharmacological treatments were administered. Supportive care included physiotherapy and developmental interventions to address hypotonia and developmental delays.

Follow-up assessments at 3, 6, and 12 months post-treatment showed remarkable improvement. At 3 months of starting treatment, neurodevelopmental milestones were appropriate for age, and the child had resumed head control and rolling over, he ran, scribbled on paper, played games and toys, made simple sentences like asking for food. By 12 months of follow up, his development was normal, with significant improvements in gross and fine motor skills, language, and social interactions. DQ improved from 50 at baseline to 100, indicating complete neurodevelopmental catch-up. These milestones were monitored using the Denver Developmental Screening Test (DDST).

A genetic counseling consultation was held with the family. Detailed information about how the condition is inherited, the risks of being carriers, and the chances of passing it on to future children. The options like carrier screening, prenatal testing, and possible reproductive choices, ensuring the parents were aware of their options moving forward. Beyond the medical details, the family was educated about the disorder itself, its symptoms and available treatments and how it might affect other relatives. Counselling also discussed the implications for other family members, including the possibility that siblings or extended relatives may also be carriers or affected.

Discussion

Biotinidase deficiency (BTD) is an autosomal recessive metabolic disorder caused by pathogenic variants in the BTD gene located on chromosome 3p25. This gene comprises four exons spanning 1,629 base pairs and encodes the enzyme biotinidase (EC 3.5.1.12), which plays a pivotal role in recycling biotin from biocytin, degraded holo-carboxylases, and protein-bound dietary sources [3]. The free biotin replenishes the intracellular pool required to activate four key carboxylases: propionyl-CoA carboxylase, β-methylcrotonyl-CoA carboxylase, pyruvate carboxylase, and acetyl-CoA carboxylase—enzymes essential for gluconeogenesis, fatty acid synthesis, and branched-chain amino acid metabolism [2, 9, 10].

BTD can manifest as either profound or partial deficiency depending on the residual enzyme activity. While symptoms most commonly arise in individuals with homozygous or compound heterozygous pathogenic variants, partial biotinidase deficiency in heterozygous individuals is typically considered asymptomatic [1, 4]. However, our case challenges this paradigm: the patient exhibited classical features of juvenile BTD despite harboring only a heterozygous mutation, suggesting that additional undetected mutations (e.g., deep intronic, structural, or regulatory variants) or tissue-specific variations in gene expression may contribute to disease manifestation [11, 12]. The identified variant, c.38_44delinsTCC, though rarely reported, may represent a functionally significant allele that warrants further study to elucidate its pathogenicity and phenotypic implications.

BTD typically presents between 2 weeks and 2 years of age, but late-onset presentations have been documented [13, 14]. The juvenile form often presents after 3 months and includes seizures, hypotonia, ataxia, alopecia, skin rashes, hearing loss, optic atrophy, and episodes of metabolic decompensation [7, 15–17]. Neurologic sequelae, including sensorineural hearing loss and developmental delays, are particularly concerning, with studies showing that 76% of untreated patients with profound BTD develop irreversible auditory impairment [18]. Interestingly, our patient exhibited prominent neurodevelopmental symptoms without any dermatological or immunological manifestations—a pattern occasionally reported but not well understood [15–17]. This phenotype may represent a milder or atypical expression of BTD or suggest variable tissue-specific vulnerability to biotin deficiency.

Diagnosis of BTD is particularly challenging in resource-limited settings like Nepal, where newborn screening (NBS), enzyme activity assays, and molecular diagnostics are often unavailable. In this case, the clinical suspicion of an inborn error of metabolism was prompted by neurodevelopmental symptoms and subsequently supported by whole exome sequencing (WES) and biochemical enzyme activity assays. WES was selected for its ability to detect coding variants efficiently and was facilitated through a research collaboration. While the sequencing revealed a heterozygous variant, the markedly reduced enzyme activity and clinical phenotype supported a diagnosis of symptomatic partial BTD. This finding supports growing evidence that monoallelic mutations may cause disease in the presence of unrecognized mutations or modifier effects [11, 12].

It is important to note that WES, while effective for identifying exonic mutations, may miss certain pathogenic variants such as large deletions, deep intronic mutations, or non-coding regulatory changes [11, 12]. Therefore, a single heterozygous mutation should not exclude the diagnosis in the presence of convincing clinical and biochemical evidence. The reliability of enzymatic assays remains high, especially when paired with genomic analysis to confirm variant pathogenicity.

Pharmacologic doses of biotin are highly effective in treating BTD, often reversing clinical symptoms and normalizing metabolic profiles. However, early initiation is crucial, as delayed therapy is associated with irreversible complications such as hearing and vision loss [19, 20]. In this case, timely administration of biotin therapy resulted in symptomatic improvement, reinforcing its therapeutic value even in partial deficiency cases. Some genotype–phenotype correlations have been suggested—for example, the p.Asp444His variant is associated with partial BTD and milder presentations [21]—but these correlations remain incompletely understood. The variant identified in this case (c.38_44delinsTCC) adds to the growing catalog of mutations in BTD and underscores the need for continued research into their clinical implications.

Global data suggest that most cases (51.5%) are identified via newborn screening, while 43.3% present symptomatically and 5.2% are diagnosed through family screening [16]. In a 40-year review of 1,113 BTD cases, multisystem involvement was common: nervous (67.2%), skin (53.7%), eyes (34.4%), auditory (26.9%), and respiratory systems (17.8%) were frequently affected, although mortality remained low (1.6%) and typically related to delayed diagnosis [16, 22]. Despite this global burden, the true epidemiology of BTD in Nepal remains unknown due to the absence of a national newborn screening program and lack of local clinical data. Our case, therefore, represents a sentinel event for raising awareness and potentially initiating future research and policy changes.

In Nepal, where advanced diagnostics remain limited and screening programs are nonexistent, clinical acumen is indispensable. A high index of suspicion for treatable metabolic conditions like BTD is necessary when encountering unexplained neurocutaneous or neurodevelopmental symptoms. Notably, all newborns referred for low dried blood spot (DBS) biotinidase activity were found to carry variants previously associated with partial biotinidase deficiency (BD), supporting the reliability of biochemical screening in identifying true positive cases [23]. The implementation of cost-effective diagnostic strategies, including dried blood spot testing and tandem mass spectrometry, alongside targeted genomic approaches, could significantly enhance early detection and intervention. Additionally, antenatal biotin supplementation may be considered in at-risk pregnancies, although more evidence is needed to support this practice [24].

In conclusion, this case not only expands the phenotypic spectrum of BTD but also illustrates the challenges of diagnosing treatable metabolic disorders in resource-limited settings. It underscores the importance of comprehensive clinical assessment, the utility of WES in ambiguous cases, and the life-altering potential of early biotin therapy. More broadly, it highlights the urgent need for newborn screening programs, local genomic research, and increased awareness among clinicians to improve outcomes for patients with rare metabolic disorders in Nepal and similar settings.

Conclusion

This case highlights the clinical and diagnostic challenges of biotinidase deficiency (BTD) in resource-limited settings like Nepal, where the absence of newborn screening and limited molecular diagnostic capacity delay timely intervention. Despite these challenges, the patient responded favorably to pharmacological doses of biotin, underscoring the efficacy of this readily available and economical therapy. However, the persistence of irreversible complications, such as hearing loss, reiterates the critical importance of early diagnosis and intervention. This report emphasizes the need for increased clinician awareness, and local research to better understand the spectrum of BTD mutations, their clinical implications and to analyze the effectiveness of implementation of newborn screening programs. Early recognition and treatment of BTD can significantly improve patient outcomes and reduce the burden of this potentially devastating yet treatable disorder.

Electronic supplementary material

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Abbreviations

BTD

Biotinidase Deficiency

WGS

Whole Genome Sequencing

WES

Whole Exome Sequencing

KCH

Kanti Children’s Hospital

GTCS

Generalised Tonic Clonic Seizure

DDST

Denver Developmental Screening Test

MCD

Multiple Carboxylase Deficiency

DBS

Dried Blood Spot

Author contributions

BK and AG conceptualized and designed the study, drafted the initial manuscript, and reviewed and revised the manuscript.ABA, BA and ST designed the data collection instruments, collected data, carried out the initial analyses, and reviewed and revised the manuscript.JK and BPM conceptualized and designed the study, coordinated and supervised data collection, and critically reviewed the manuscript for important intellectual content.All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work.

Funding

No financial or non-financial benefits have been received or will be received from any party related directly or indirectly to the subject of this article.

Data availability

The authors confirm that the data supporting the findings of this study are available within the article and supplementary files.

Declarations

Ethical approval

Case reports are exempt from ethical approval in our institution, Kanti Children’s Hospital, Maharajgunj, Kathmandu.

Consent to participate and publication

Written informed consent was obtained from the patient’s parents for participation and publication of this case reports and accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal on request.

Competing interests

The authors declare no competing interests.