|Ahead of print publication
Prevalence of subclinical Vitamin K deficiency in early infancy in exclusively breast-fed term infants
Gitanjali Jain1, KM Adhikari2, Gautam Kumar Vasnik3, Daljit Singh4, Venkatesan Somasundaram5, Rakesh Gupta6, Puja Dudeja7, Subhash Chandra Shaw4
1 Department of Pediatrics, AIIMS, New Delhi, India
2 Department of Pediatrics, Armed Forces Medical College, Pune, Maharashtra, India
3 Department of Pathology, Base Hospital, Delhi, Cantt, India
4 Department of Pediatrics, Army Hospital (R and R), Delhi, India
5 Department of Pathology, Army Hospital (R and R), Delhi, India
6 Department of Pediatrics, Government Institute of Medical Sciences, Noida, Uttar Pradesh, India
7 Department of Community Medicine, Army College of Medical Sciences, Delhi, Cantt, India
|Date of Submission||12-May-2022|
|Date of Decision||06-Jul-2022|
|Date of Acceptance||29-Aug-2022|
|Date of Web Publication||11-Oct-2022|
Subhash Chandra Shaw,
Department of Pediatrics, Army Hospital (R and R), Delhi - 110 010
Source of Support: None, Conflict of Interest: None
Background: Late Vitamin K deficiency bleeding, is a disease of exclusively breast-fed infants attributable to poor content of Vitamin K in breast milk. We aimed to estimate the prevalence of subclinical Vitamin K deficiency, at 3 months of age in exclusively breast-fed term infants who were administered routine Vitamin K 1 mg intramuscular at birth. Methods: This prospective observational study was conducted between June 2018 and June 2020 in a Tertiary Care Teaching Hospital in Western India. Consecutive inborn infants, having received Injection Vitamin K at birth and on exclusive breast feeds attending immunization at 3 months of age were included. Half milliliter of venous blood was withdrawn for the estimation of Proteins Induced by Vitamin K Absence or Antagonist-II (PIVKA-II). Subclinical Vitamin K deficiency was defined as PIVKA-II level of more than 2 ng/mL. Results: The mean age of infants was 3.2 ± 0.2 months and the PIVKA II levels ranged from 0.19 to 10.07 ng/ml. Subclinical Vitamin K deficiency was present in 62 (32.63%) out of 190 infants. Conclusion: Significantly raised PIVKA II levels (>2 ng/mL) were detectable in 32.6% of infants after 3 months of age who had received 1 mg of Vitamin K at birth.
Keywords: Proteins induced by Vitamin K absence or antagonist-II, subclinical Vitamin K deficiency, Vitamin K deficiency bleeding
|How to cite this URL:|
Jain G, Adhikari K M, Vasnik GK, Singh D, Somasundaram V, Gupta R, Dudeja P, Shaw SC. Prevalence of subclinical Vitamin K deficiency in early infancy in exclusively breast-fed term infants. J Mar Med Soc [Epub ahead of print] [cited 2022 Dec 7]. Available from: https://www.marinemedicalsociety.in/preprintarticle.asp?id=358298
| Introduction|| |
Vitamin K deficiency bleeding (VKDB) is classified as per age of presentation of bleed, as early (within 24 h), classical (1–7 days after birth), and late (after 1 week till 6 months of life). Late VKDB, the most dangerous of all as it frequently results in intracranial bleed, and is a disease of exclusively breast-fed infants (attributable to poor content of Vitamin K in breast milk). The incidence of VKDB in late Vitamin K deficiency ranges from 1 in 15,000 to 1 in 20,000 births, is seen mainly in exclusively breast-fed infants and with associated risk factors. In subclinical Vitamin K deficiency, although there is no clinical bleed, hepatic stores of Vitamin K are not adequate to ensure complete gamma-glutamyl carboxylation of the Vitamin K-dependent procoagulant proteins, Factor II, VII, IX, and X (Vitamin K-dependent coagulation factors). Thus, in states of Vitamin K insufficiency, functionally defective undercarboxylated factors are being released into the circulation and these factors are collectively termed Proteins Induced by Vitamin K absence or antagonist-II (PIVKA-II)., We aimed to estimate prevalence of subclinical Vitamin K deficiency, at 3 months of age in exclusively breast-fed term infants who were recipient of routine Vitamin K 1 mg intramuscular at birth. The secondary objective was to estimate prevalence of VKDB, by 3 months of age.
| Methods|| |
This prospective observational study was conducted between June 2018 and June 2020 in a tertiary care teaching hospital in western India. Informed written consent was obtained from one of the parents. The study was approved by the institutional ethics committee. Consecutive infants who were born at term in the same hospital received injection of Vitamin K at birth as per records and on exclusive breast feeds, who attended pediatric outpatient department for immunization at 3 months of age were included in this study. The infants who had received antibiotics for more than 7 days during the first 3 months of life or had features of direct hyperbilirubinemia were excluded. Neonates whose mothers were taking drugs that interfere with Vitamin K function (anticonvulsants, antibiotics, Vitamin K antagonists) were also excluded from the study.
Details of documented or suspect VKDB since birth and other antenatal, perinatal, and postnatal information were recorded in the pro forma. After taking informed consent, 0.5 ml of venous blood was drawn into plain vacutainers for estimation of PIVKA-II. Samples were allowed to clot and then were kept at 2°C–8°C till overnight before centrifugation for 15 min at 1000 ×g. The supernatants were stored at −80°C and processed for the estimation of PIVKA-II.
The sample size was estimated based on a reported incidence of 12.69% of infants having detectable PIVKA-II at 3 months of age as studied by Cornelissen et al., To detect this proportion in a limited population setting, a sample size of 157 was calculated. To cater for 20% of samples collected but not processed or hemolyzed, a minimum sample size of 188 infants was arrived at.
PIVKA-II levels were estimated by sandwich ELISA technique using the Human PIVKA-II kits, Elabscience®, sensitivity of the test kit being 0.10 ng/ml, detection range from 0.16 to 10 ng/ml. This ELISA kit applies to the in vitro quantitative determination of Human PIVKA-II concentrations in serum or plasma. 100 μL of sample was processed for optic density quantification as per the manufacturer's guidelines which were measured at a wavelength of 450 ± 2 nm spectrophotometrically. The optical density value was proportional to the Human PIVKA-II concentration. PIVKA-II level of more than or equal to 2 ng/mL was considered “detectable.”
| Results|| |
Among the enrolled 198 infants attending the immunization clinics at 14 weeks, 8 samples got hemolyzed and could not be processed for PIVKA-II levels. Out of the rest 190 infants, 88 (46.3%) were female and 102 (53.7%) were male. One hundred and thirty-six (71.6%) neonates were born vaginally and 54 (28.4%) neonates were born by cesarean section. Eight-five (44.7%) neonates were born to primipara mothers and 105 (55.3%) were born to multipara mothers. Ninety-eight (51.6%) mothers took an exclusively vegetarian diet and 92 (48.4%) mothers took nonvegetarian/mixed diet. One hundred and seventy-one (90%) mothers took green vegetables in diet at least 3 to 4 times/week.
At the time of enrolment, the mean age of infants was 3.2 ± 0.2 months and the mean age of mothers was 25.3 ± 3.7 years. The PIVKA II levels ranged overall from 0.19 to 10.07 ng/ml. For the cutoff value for Vitamin K deficiency, PIVKA-II level was taken as more than 2, and >2 ng/mL of PIVKA-II was present in 32.63% (62/190) infants. The association of various maternal and neonatal factors with Vitamin K deficiency is depicted in [Table 1]. None of the maternal factors such as parity, diet, and mode of delivery had any association with PIVKA-II values in the infants. None of the enrolled neonates had any history or documentation of any bleeding manifestations till the time of inclusion in the study.
|Table 1: Association of maternal and neonatal factors with Vitamin K deficiency (proteins induced by Vitamin K absence or antagonist-II: >2 ng/ml)|
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| Discussion|| |
The current study was aimed at only to find the prevalence of subclinical Vitamin K deficiency at 3 months of age by estimating the level of PIVKA-II. We state that, though not a novel or complex study, finding 32.63% of infants having PIVKA-II values above cutoff level is a significant observation. The prevalence of late VKDB is about 35/100,000 live births in the infants who did not receive Vitamin K prophylaxis at birth. A large study published in the Lancet found a five-fold reduction in mortality from hemorrhage in the infants who were administered 1 mg of Vitamin K3 (menadione) at delivery. We continue to encounter in our clinical practice, the cases of late VKDB in the infants even though received Vitamin K at birth. In subclinical Vitamin K deficiency, though there is no clinical bleed, hepatic stores of Vitamin K are not sufficient to ensure full gamma-glutamyl carboxylation of the Vitamin K-dependent procoagulant proteins, factor II, VII, IX, and X. At present, PIVKA is considered a more sensitive indicator than the prothombin time as a marker of Vitamin K deficiency. Although our center has a robust Vitamin K injection practice with mandatory documentation, we decided to look for possible subclinical Vitamin K deficiency despite receiving the supplement.
Most of the studies in past have measured PIVKA-II levels in the early neonatal period, soon after injectable/oral Vitamin K prophylaxis. Motohara et al. detected PIVKA-II in cord blood in 21.5% of 102 samples. However, among those who received Vitamin K at birth, only 11%–18% were PIVKA-II positive at 3–5 days and their PIVKA-II levels had remained unchanged since birth. Cornelissen et al. demonstrated detectable PIVKA-II in 11.5% of infants at 3 months of age whereas Widdershoven et al. detected PIVKA II in 7.5% of breast-fed infants at 3 months of age. The higher levels of PIVKA II in our study as compared to earlier studies, might be due to the greater sensitivity of the latest ELISA kits that we used. However, none of the infants in our study had any clinical bleeding manifestations despite detectable PIVKA II levels. Our findings were similar to a study done by Chuansumrit et al., in Thai neonates. In our study too, none of the maternal factors such as age, parity, socioeconomic status, or diet had any correlation with PIVKA-II levels in the infants.
The strength of our study is that all the infants included were exclusively breastfed, and the measurement of serum PIVKA II levels was combined with 14 weeks' immunization, thus avoiding any extra hospital visits of the infants for the study. The ELISA kits that we used for measuring PIVKA II levels had a high sensitivity (up to 0.10 ng/ml) as well. We would like to highlight the limitation of the sample size. Calculation based on the Poisson regression model, the ideal method of sample size calculation for research of this nature in a population-based study, would have mandated the inclusion of a higher number of infants. This was not feasible in the intended study framework and logistically.
| Conclusion|| |
We conclude that notably raised PIVKA II levels (>2 ng/ml) were detectable in 32.6% of infants after 3 months of age suggesting biochemical Vitamin K deficiency, in exclusively breast-fed infants who were born at term and had received 1 mg of Vitamin K at birth. None of the infants had any clinical manifestations of late VKDB. The role of an additional dose of Vitamin K in reducing the subclinical Vitamin K deficiency needs to be studied further in a larger series.
This article is based on the Armed Forces Medical Research Council project 5032/2018.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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