Nonketotic Hyperglycinemia Diagnosis

A Typical Neonatal NKH Diagnosis

Below is a fictional, but fairly typical experience of being diagnosed. It’s a scary time, so if you’ve recently been diagnosed we send our love.

Birth and Initial Presentation

  • A baby is born at term, after an uneventful pregnancy and delivery.
  • Baby is released home, and the parents notice the baby is increasingly lethargic and not interested in feeding.
  • They visit their paediatrician who refers them to the emergency department at their local hospital.

Initial Hospital Admission

  • Baby presents with severe lethargy with minimal response to painful stimuli and gross hypotonia.
  • They do a sepsis work up (including blood and urine cultures and a lumbar puncture).
  • Baby is started wide spectrum antibiotics for sepsis.
  • An echocardiogram (sonogram of the heart) is done, as well as a CT but both are unremarkable

Admission to NICU

  • The baby’s breathing is shallow, resulting in respiratory acidosis (where decreased breathing increases the amount of carbon dioxide in the blood, and decreases the blood’s pH – called acidosis). To prevent a build up of CO2, the baby is put on mechanical ventilation.
  • The baby is declared nill by mouth (NPO) and placed on a intravenous dextrose infusion (dextrose is a sugar that your body produces naturally. In this case it’s used as a food replacement to minimise any stress on the body)
  • The baby is losing it’s baby reflexes.
  • A Crainial Ultrasound was normal on the left and right.

Seizures Start + Neuro Tests

  • Clinical seizures appear on Day 5 of life, and are treated with a loading dose of Phenobarbital.
  • An EEG was done to view brain activity, and a suppression burst pattern with abnormal activity was shown.
  • An ultrasound was done on the liver, returned normal
  • Urine, serum amnio and organic acids, lactate acids and pyruvate levels were done and all within normal limits.
  • A muscle/skin biopsy is done. Returns normal.
  • An MRI is done and showed a thin, short corpus collosum, increased T2 signal in the white matter (especially frontally), absent myeline in the PLIC, with abnormal signalling in the brain stem tracts.
  • A second lumbar puncture is done, and CSF and plasma are sent for amino acids, and returns high glycine levels.

Diagnosis and Discharge

  • A diagnosis of Nonketotic Hyperglycinemia was made from the MRI and the elevated glycine ratio between the CSF and plasma.
  • The baby was started on Sodium Benzoate and Dextromethorphan.
  • Baby was weaned from the mechanical ventilation and over the next week showed improved tone, returned reflexes and was able to feed orally.
  • Baby was discharged at 25 days of life with medication, a protein restricted diet and hospice care

Prenatal Diagnosis

In the 1980s, measurement of amniotic fluid glycine levels and the ratio between glycine and serine in amniotic fluid was used in some cases to detect fetuses affected with NKH.

There were, however, instances in which control values and values in affected fetuses overlapped. This method of diagnosis was abandoned with the advent of chorionic villus sampling (CVS) biopsy.

Biochemical diagnosis of NKH by CVS is performed at eight to 12 weeks gestation to measure GCS activity in the trophoblast. Parents considering
prenatal CVS GCS testing should be informed that there is an incidence of 1 percent false positive and 1 percent false negative results associated with this procedure.

The false negative results occur secondary to residual GCS activity in affected fetuses. Parents should also be counseled in the risk to the fetus associated with CVS, including limb and oromandibular defects and fetal loss.

In addition to biochemical diagnosis, genotype determination can be performed both prenatally using DNA extracted from fetal cells via amniocentesis or CVS, or postnatally.

There are three genes that code for the GCS (GLDC, AMT, and GCSH) on different chromosomes. Because no common mutations have been identified, full sequencing of these three genes must be performed, making DNA analysis an expensive and lengthy process.

Researchers are trying to develop screening tools for the three affected genes to improve the time and cost effectiveness of genetic testing. Conter and colleagues performed mutation analysis of the genes coding for the GCS complex using denaturing high-performance liquid chromatography (DHPLC) in 28 patients with NKH with a resulting sensitivity and specificity of 97 percent and 96 percent, respectively.

Genetic testing combined with GCS activity analysis can improve the reliability of prenatal diagnosis.

Postnatal Diagnosis

Prompt diagnosis of an inborn error of metabolism is possible if appropriate diagnostic testing is ordered. An inborn error of metabolism should be suspected in any newborn presenting with rapid clinical deterioration that does not respond to symptomatic therapy following a normal delivery
and symptom-free newborn period.

A routine workup for an infant with a suspected inborn error of metabolism should include a complete blood cell count; an arterial blood gas; an electrolyte panel; blood glucose; liver function tests; and serum ammonia, lactate, glycine, and pyruvate levels.

A urine sample should be sent for ketones, reducing substances, and urine amino and organic acid screens. A lumbar puncture should be performed and a specimen sent for CSF amino acids. These samples should be collected prior
to stopping feedings or beginning specific treatment to avoid
obtaining false negative results.

The “biochemical hallmarks” of NKH are elevated plasma and CSF glycine concentrations, leading to an elevation of the CSF to plasma glycine ratio above normal (,0.04).

In infants whose presentation suggests NKH, plasma and CSF glycine levels should be drawn simultaneously to determine the CSF to plasma glycine ratio. A CSF to serum glycine ratio of .0.08, from concurrently drawn samples, is considered diagnostic for typical or neonatal-onset NKH.

This method of diagnosis does not distinguish transient cases of NKH in which the CSF to plasma glycine ratio is initially elevated but subsequently normalizes. Practitioners need to be aware that CSF contaminated with blood is not a valuable specimen. The finding of hyperglycinemia in CSF may be the result of a bloodstained spinal tap. The presence of erythrocytes in CSF falsely elevates amino acid concentrations.

A global elevation of amino acids in the CSF should alert the provider to the possibility of a blood-contaminated specimen. CSF glycine levels from an overtly bloodstained spinal tap should not be used for the diagnosis
of NKH.

The “gold standard” for the confirmation of a diagnosi of NKH, first described by Hayasaka and colleagues in 1990, is a liver glycine cleavage complex and component assay. Unfortunately, obtaining the large liver sample required for diagnosis (.80 mg of tissue) is difficult, particularly in
a sick neonate, and, therefore, is rarely done.

In a recent survey of families registered with International NKH Family
Network, Hoover-Fong and associates found that only nine out of 65 respondents had a diagnosis confirmed by liver GCS assay. Diagnosis by liver GCS assay effectively rules out cases of transient NKH, as there are no reported cases of infants with complete absence of liver GCS activity who later recovered.

Tandem mass spectrometry is used in newborn screening for inborn errors of metabolism including disorders of metabolism of the urea cycle, amino acids, organic acids, and fatty acid oxidation. Tandem mass spectrometry
can also be used to measure glycine levels in the routine newborn screening blood spot samples; however, the positive predictive value of the newborn glycine level for NKH is 5.7 percent, with a high false negative rate (78 percent).

In a study by Tan and colleagues of 624,000 babies that were screened with tandem mass spectrometry, only 35 had an isolated increase in glycine for which repeat specimens were sent. 23 Of those 35 infants, only two were subsequently confirmed to have NKH. In both of these two cases, the infants presented with severe forms of classical NKH, and diagnosis with plasma and CSF glycine levels was made before the newborn screening results were returned, leading the researchers to conclude that the results of newborn screens have no impact on the clinical management of patients with

In addition, although NKH is included on some state screens, it is not a part of the Recommended Uniform Screening Panel for all states. In 2006, Kure and colleagues described using a glycine breath test to diagnose NKH. Carbon dioxide is a byproduct of the metabolism of glycine by the GCS. The researchers identified lower exhaled carbon dioxide levels in five patients
with NKH who were given a test load of 10 mg/kg of glycine versus control subjects administered the same glycine load. If further research with a larger patient population verifies the prognostic predictive value of the glycine breath test, it would provide the least invasive means of diagnosis
for NKH.

Differenital Diagnosis of Nonketotic Hyperglycinemia

Inborn Errors of Metabolism

The most significant differential diagnoses for NKH are the inborn errors of metabolism that lead to ketotic hyperglycinemia: propionic, methylmalonic, and isovaleric acidemias. These inborn errors of metabolism lead to accumulation of branched-chain amino acid metabolites, which suppress the hepatic GCS, causing hyperglycinemia.

In ketotic hyperglycinemia, however, GCS activity of the CNS is not inhibited, accounting for CSF glycine levels and a CSF to plasma glycine ratio within normal limits. Analysis of urine organic acids in ketotic hyperglycinemia will demonstrate ketonuria and specific organic aciduria.


The symptoms associated with NKH in the neonatal period, including feeding difficulties, apnea, hypotonia, and lethargy, may be nonspecific and mimic signs of neonatal sepsis. For this reason, a full sepsis workup including blood, urine, and CSF cultures should be obtained, and antibiotics
and antivirals should be administered until NKH is diagnosed or pending negative bacterial and viral cultures.

Acute neurologic disorders leading to neonatal encephalopathy, such as hypoxic-ischemic encephalopathy, congenital stroke, and meningitis, may present with the same clinical symptoms as NKH. These disorders may also be associated with transient elevations in the CSF to plasma glycine ratio secondary to impairment of glycine metabolism in the brain during periods of encephalopathy.25 A thorough delivery history will help differentiate NKH from traumatic brain injury in many cases.

Credit Side note: Most of this is taken from the research paper Neonatal Nonketotic Hyperglycinemia: A Case Study and Review of Management for the Advanced Practice Nurse by Joanna L. Mulligan