The aim of this study is to assess masseter muscle thickness using ultrasounds during oral feeding process in premature infants with gestational age from 28+0/7 to 32+0/7 weeks, taking into account the qualitative and quantitative aspects of oral feeding. The main question it aims to answer is: Does the thickness of the masseter muscle influence the course of development of the feeding function in premature newborns? The study will assess the influence of masseter muscle thickness on the oral feeding process in premature infants, which has significant cognitive value due to the dynamic development of feeding function in this group of newborns. Transitioning from enteral feeding via a nasogastric tube to full oral feeding is a crucial stage in the development of one of the most fundamental life functions - nutrition. Complete, effective, and safe oral feeding is also a factor in determining a newborn's discharge from the hospital. Research in the area of feeding is an important and ongoing goal for improving neonatal care in neonatal units. Furthermore, this study will provide preliminary data on masseter muscle thickness in premature infants, which may serve as a basis for further research.

The study will include premature infants who meet the inclusion and exclusion criteria. The first ultrasound examination (USG) of the masseter muscle thickness will be performed between 7 and 10 days of life. The second examination will be performed at the initial stage of oral feeding - when the child is consuming orally 25 to 50% of its daily nutritional requirements. If the daily requirement is expressed as a volume range (e.g., 40-45ml/serving, i.e., 320ml-360ml/day), the lower expected value will be used to calculate the child's food intake. The third examination will be performed when the child has achieved full oral feeding. The ultrasound examination will include measurement of the masseter muscle in its thickest section, from the external to the internal fascia (including their thickness), in longitudinal and transverse sections, using a linear transducer designed for musculoskeletal examinations. The transducer will be placed perpendicular to the mandibular ramus. Muscle thickness is defined as the maximum distance between the external and internal fascia. The newborn will be positioned in a supinated position during the examination. During the ultrasound examination, routine procedures will be used to support the newborn's self-regulation (e.g., swaddling, ensuring the ability to clasp hands or hold them near the face, ensuring a quiet environment, avoiding excessive and sudden sounds, etc.). A water-based transmission gel, heated in an ultrasound gel warmer to 36 degrees Celsius (98 degrees Fahrenheit), will be used during the examination to ensure the newborn's comfort and consistent examination conditions. During the examination, the ultrasound transducer will be placed on the left and then the right side of the baby's face to assess the left and right masseter muscles. The examination will be performed by a single person - a neonatologist and pediatrician specialist with extensive experience in performing ultrasound imaging in premature newborns. Performing the examination by a single person will ensure consistent pressure from the ultrasound transducer on the baby's tissues. The examination will be recorded on an ultrasound machine - recording time: 10 seconds for each area examined, i.e., a total of 20 seconds of recording (left and right masseter muscles). Data will be transferred from the ultrasound machine using a portable storage device (flash drive). The recorded examinations will then be evaluated by a radiology and imaging specialist. Masseter thickness will be measured when the muscle relaxes, defined as the absence of sucking movements in the child.

Within a maximum of 3 days from the day on which the second and third ultrasound examinations are performed, study will also be carried out to assess the course of the feeding session to evaluate the quantitative and qualitative aspects of oral feeding. Newborns will be fed in the side-lying position, positioned on their left side. The examination will include assessment of the baby before, during, and after feeding:

• 2 minutes before feeding, oxygen saturation (SpO2) and heart rate (HR) + the level of activity according to the Neonatal Behavioral Assessment Scale (NBAS);
• 3 minutes into feeding, SpO2 and HR + the level of activity according to the NBAS;
• at the end of feeding, SpO2 and HR + the level of activity according to the NBAS + volume of food consumed;
• 10 minutes after feeding, SpO2 and HR + the level of activity according to the NBAS.

SpO2 and HR will be measured using a pulse oximeter (Masimo Rad97). The pulse oximeter sensor will be placed on the right lower limb. Feeding time: measured from the moment the teat is placed in the baby's mouth until the end of feeding. The total feeding time does not include breaks in feeding, such as burping or any interruption caused by a choking episode due to sucking, breathing, and swallowing discoordination. Maximum feeding session time: 30 minutes - measured from the moment the baby is taken from the bed until the end of the feeding (removal of the teat from the baby's mouth). Breaks in feeding, such as burping, are included in the total feeding session time. After feeding, the baby may be placed in the bed or placed on the mother's or father's chest for kangaroo care. Both the feeding time and the total feeding session time will be recorded. The occurrence of choking episodes and the total duration of SpO2 drops will also be taken into account in the study. If the use of extraoral aids to improve feeding efficiency is necessary, this will be noted in the study protocol. These aids improve the seal of the lips around the teat in active newborns during feeding. They are used only when necessary, i.e., if a still-active baby develops lip slack during feeding, resulting in decreased feeding effectiveness. The volume of food consumed will be expressed as a percentage of the expected volume of food consumed per serving. The study will be video recorded. Description of feeding position: The newborn is placed in a lateral position on the examiner's lap, on the left side, at a 30-45° angle (with a folded blanket/pillow or footrest between the knees and the newborn). The baby's head is symmetrically positioned between the shoulders, supported by the hand of the person feeding the baby. The shoulder girdle is higher than the pelvic girdle, and the head and back are in a straight line - slight, natural body flexion is permitted. The legs are bent at the hips at an angle of up to 90°. The knees and ankles are in natural flexion. For the purposes of the study, data will be collected regarding the baby's maturity on the day of delivery, the type of delivery, sex, and the baby's weight at delivery. Furthermore, on the days when the ultrasound examination will be performed, the newborn's anthropometric data will be collected: body weight and head circumference. To avoid exposing newborns to additional procedures related to obtaining the above data, the baby's body weight will be recorded from the routine daily measurement, and head circumference from the last routine weekly measurement. Variables related to the process of introducing full oral feeding will also be analyzed: the day of life in which the baby reaches 50% oral feeding; the number of days from the first feeding attempt to the introduction of full oral feeding; and the average daily weight gain calculated from the first feeding attempt to the introduction of full oral feeding, as well as between the first and second, and second and third, masseter ultrasound examinations. Examination procedure: Examination I: Bilateral ultrasound assessment of the masseter muscle on the 7th-10th day of life; body weight + head circumference; Examination II: Bilateral ultrasound assessment of the masseter muscle when the infant is orally consuming 25-50% of its daily requirement; body weight + head circumference; Feeding session assessment (SpO2; drops in SpO2 below 85%, HR; feeding duration; feeding session duration; need for extraoral support; occurrence of choking episodes, activity status according to the NBAS scale; amount of food consumed); Examination III: bilateral ultrasound assessment of the masseter muscle when the child is orally consuming 100% of its daily requirement; body weight + head circumference; feeding session assessment (SpO2; drops in SpO2 below 85%, HR; feeding duration; feeding session duration; need for extraoral support; occurrence of choking episodes, activity status according to the NBAS scale; amount of food consumed).

Inclusion Criteria:

• 28+0/7-32+0/7 weeks of gestational age;
• use of one specific type of teat;
• passive oxygen therapy is permitted;
• written informed consent obtained from parents.

Exclusion Criteria:

• newborns with defects that may significantly affect the development of feeding function such as: cleft lip and/or palate, facial paralysis, and other craniofacial defects;
• newborns with a shortened anterior lingual frenulum according to the E. Coryllos classification - meeting the criteria for frenotomy according to the Hazelbaker/Amir scale;
• detected genetic defects, metabolic diseases, or suspected cases of their presence;
• newborns requiring intubation up to 10 days of life;
• low Apgar score (less than 5 points at 5 and 10 minutes of life);
• newborns with visible neurological disorders (e.g. with symptoms of hypertonia, impaired reflex reactions in the orofacial area) and/or significant abnormalities in central nervous system (CNS) imaging studies (e.g. III-IV degree intraventricular hemorrhages, CNS defects, hydrocephalus);
• newborns of mothers with anhydramnios;
• lack of consent to conduct at least two bottle-feeding sessions;
• a child fed with a different type of teat than the one used in the study.

The study will include premature newborns hospitalized in the Neonatology, Intensive Care and Neonatal Pathology Clinic, Polish Mother's Memorial Hospital Research Institute, Łódź, Poland