Development of Personalized Nutrition: Applications in Lactose Intolerance Diagnosis and Management.

Recent discoveries in the "omics" field and the growing focus on preventive health have opened new avenues for personalized nutrition (PN), which is becoming an important theme in the strategic plans of organizations that are active in healthcare, food, and nutrition research. PN holds great potential for individual health optimization, disease management, public health interventions, and product innovation. However, there are still multiple challenges to overcome before PN can be truly embraced by the public and healthcare stakeholders. The diagnosis and management of lactose intolerance (LI), a common condition with a strong inter-individual component, is explored as an interesting example for the potential role of these technologies and the challenges of PN. From the development of genetic and metabolomic LI diagnostic tests that can be carried out in the home, to advances in the understanding of LI pathology and individualized treatment optimization, PN in LI care has shown substantial progress. However, there are still many research gaps to address, including the understanding of epigenetic regulation of lactase expression and how lactose is metabolized by the gut microbiota, in order to achieve better LI detection and effective therapeutic interventions to reverse the potential health consequences of LI.

Lactose intolerance: An update on its pathogenesis, diagnosis, and treatment.

Lactose intolerance has a high prevalence worldwide, ranging between 57% and 65%. It is caused by a reduction or loss of the activity of the intestinal enzyme lactase-phlorizin hydrolase, responsible for the digestion of lactose. This alteration determines an increased osmotic load in the small intestine and the fermentation of lactose by the bacterial flora, which leads to a high production of short-chain fatty acids and gas. This is followed by the onset of abdominal pain, diarrhea, and flatulence. In addition to these problems, it was found that subjects with lactose intolerance have an increased risk of developing various extra-intestinal diseases, including cancers. The diagnosis is essential to undertake an adequate treatment and, for this purpose, different methods have been tested. These include genetic test, hydrogen breath test (HBT), quick lactase test, and lactose tolerance test. HBT is the most used method because it is non-invasive, inexpensive, and highly sensitive and specific, as well as easy to perform. In clinical practice, the other methods are mainly used as HBT integration tests. There are also many therapeutic options. An appropriate intervention concerns the dietetic style, such as the consumption of lactose-free foods, but with nutritional characteristics comparable to dairy products. Other valid choices are represented by the use of exogenous enzymes, probiotics, prebiotics, the selection of milk containing specific types of beta-caseins. This review is intended to illustrate the diagnostic methods currently available and the possible therapeutic options for lactose intolerance.

Milk Containing A2 ß-Casein ONLY, as a Single Meal, Causes Fewer Symptoms of Lactose Intolerance than Milk Containing A1 and A2 ß-Caseins in Subjects with Lactose Maldigestion and Intolerance: A Randomized, Double-Blind, Crossover Trial.

Acute-feeding and multiple-day studies have demonstrated that milk containing A2 ß-casein only causes fewer symptoms of lactose intolerance (LI) than milk containing both A1 and A2 ß-caseins. We conducted a single-meal study to evaluate the gastrointestinal (GI) tolerance of milk containing different concentrations of A1 and A2 ß-casein proteins. This was a randomized, double-blind, crossover trial in 25 LI subjects with maldigestion and an additional eight lactose maldigesters who did not meet the QLCSS criteria. Subjects received each of four types of milk (milk containing A2 ß-casein protein only, Jersey milk, conventional milk, and lactose-free milk) after overnight fasting. Symptoms of GI intolerance and breath hydrogen concentrations were analyzed for 6 h after ingestion of each type of milk. In an analysis of the 25 LI subjects, total symptom score for abdominal pain was lower following consumption of milk containing A2 ß-casein only, compared with conventional milk (p = 0.004). Post hoc analysis with lactose maldigesters revealed statistically significantly improved symptom scores (p = 0.04) and lower hydrogen production (p = 0.04) following consumption of milk containing A2 ß-casein only compared with conventional milk. Consumption of milk containing A2 ß-casein only is associated with fewer GI symptoms than consumption of conventional milk in lactose maldigesters.

Lactose Intolerance: A Concise Review to Skim the Surface.

Lactose intolerance is a common but poorly understood cause of gastrointestinal symptoms. Contrary to popular belief, there is much more to its diagnosis beyond symptoms with exposure and management beyond milk- and dairy-product avoidance. In this article, we review definitions, genetic basis, pathogenesis, clinical signs, as well as diagnostic and management strategies.

Comparison of the impact of bovine milk ß-casein variants on digestive comfort in females self-reporting dairy intolerance: a randomized controlled trial.

BACKGROUND: Lactose malabsorption (LM) is a major cause of digestive discomfort from dairy products. Recently, a role for bovine ß-casein A1 has been proposed. OBJECTIVES: We examined whether there are distinct symptoms of digestive discomfort due to either lactose or differing bovine ß-casein types. METHODS: Women (n = 40; age: 25.2 ± 0.5 y) with self-reported varying dairy tolerance underwent a 50-g lactose challenge. Based on postchallenge LM and digestive discomfort, participants were classified as either lactose intolerant (LI; n = 10, self-reported intolerant, diagnosed lactose intolerant), nonlactose dairy intolerant (NLDI; n = 20, self-reported intolerant, diagnosed lactose tolerant), or dairy tolerant (DT; n = 10, self-reported tolerant, diagnosed lactose tolerant). In a double-blinded randomized sequence, participants consumed 750 mL conventional milk (CON; containing A1 and A2 ß-casein and lactose), a2 Milk (A2M; exclusively containing A2 ß-casein with lactose), or lactose-free conventional milk (LF-CON; containing A1 and A2 ß-casein without lactose). Subjective digestive symptoms and breath hydrogen (measuring LM) were recorded regularly over 3 h, and further ad hoc digestive symptoms over 12 h. RESULTS: LI subjects experienced prolonged digestive discomfort with CON milk. A2M reduced (P < 0.05) some symptoms (nausea: A2M 8 ± 3 mm compared with CON 15 ± 3mm; fecal urgency: A2M 4 ± 1 compared with CON 10 ± 3 mm), and attenuated the rise in breath hydrogen over 3 h, relative to CON milk (A2M 59 ± 23 compared with CON 98 ± 25 ppm at 150 min; P < 0.01). In contrast, NLDI subjects experienced rapid-onset, transient symptoms (abdominal distension, bloating, and flatulence) without increased breath hydrogen, irrespective of milk type. CONCLUSIONS: In LI individuals, LM and digestive comfort with lactose-containing milks was improved with milk containing exclusively A2 ß-casein. Furthermore, self-reported dairy intolerance without LM (NLDI) is characterized by early-onset digestive discomfort following milk ingestion, irrespective of lactose content or ß-casein type. This trial was registered at www.anzctr.org.au as ACTRN12616001694404.

Does primary lactase deficiency reduce bone mineral density in postmenopausal women? A systematic review and meta-analysis.

Postmenopausal osteoporosis is a significant cause of morbidity and mortality. The role of primary lactase deficiency (PLD) in its development is not clear. This meta-analysis showed that PLD is a risk factor for osteoporosis in postmenopausal women. These women need special attention in terms of screening for osteoporosis and its prevention. INTRODUCTION: Postmenopausal osteoporosis is an important predictor of bone fractures. The purpose of the study was to conduct a systematic review and meta-analysis of association of PLD and bone mineral density (BMD) in postmenopausal women. METHODS: The electronic databases PubMed, Scopus, and Web of Science were searched over the course of July 2017 for any date of publication without language limitation. Studies were included in the meta-analysis if the diagnosis of PLD was made by genetic testing or H-2 breath tests and the diagnosis of osteoporosis was made by a modern reliable method for BMD measurement. Two investigators conducted a comprehensive, independent review of all the papers. Five of the studies initially identified met the inclusion criteria. We used MOOSE guidelines for abstracting data and assessing data quality and validity. Meta-analysis was performed using the random effects model. RESULTS: Five case-control studies with 2223 participants and 763 lactase-deficient cases fulfilled the inclusion criteria. Meta-analysis showed a significantly higher bone density Z-score in absorbers (mean difference 0.20, CI (0.14-0.27), P = 0.000), with no significant heterogeneity among the studies. Moreover, the Z-score in the vast majority of the measured sites (femoral head, femoral neck, lumbar spine, radius, and Ward's triangle) was significantly higher in absorbers. There was no significant overall difference in BMD in g/cm2 between absorbers and non-absorbers, but a significantly higher BMD using g/cm2 was observed in absorbers in the total hip site. CONCLUSIONS: Postmenopausal women with PLD had lower Z-scores at most anatomic sites compared to healthy controls.

Gastrointestinal Symptoms and FODMAP Intake of Aged-Care Residents from Christchurch, New Zealand.

Studies on fermentable oligo-, di-, and monosaccharides as well as polyols (FODMAPs) intake in older adults are lacking. This study investigated the relationship between gastrointestinal (GI) symptoms and FODMAPs in aged care residents. The Gastrointestinal Symptom Rating Score questionnaire modified for patients with IBS (GSRS-IBS) was used to identify participants with IBS-like symptoms. Dietary intake was assessed for a subgroup of participants with highest total GSRS-IBS score (symptomatic cases) and age, sex, and level of care matched participants with low total GSRS-IBS score (asymptomatic controls). Seventy-four participants with a mean (SD) age of 86 (6.6) years completed the GSRS-IBS questionnaire and dietary data were collected using food diaries from a subsample of 27 symptomatic and 27 asymptomatic participants. The study found many older adults with functional gut symptoms. There were no differences between the groups for FODMAP intake and no significant relationship was found between FODMAP intake and total GSRS-IBS score. Lactose from milk and milk-based desserts was the biggest FODMAP contributor (16 g/day) and a significant relationship between total FODMAP intake and diarrhoea was found. A larger study sample in future studies is required to better capture symptomatic cases and manipulation of dietary FODMAPs may assist with the management of IBS in the elderly.

Combination of real-time PCR and sequencing to detect multiple clinically relevant genetic variations in the lactase gene.

BACKGROUND: Lactase persistence is an autosomal dominant trait commonly distributed in Europe as well as some parts of east Africa and the Arabian Peninsula. Using real-time PCR to detect the -13910C > T variant common in the European population is a reliable analysis although other variants in the probe-binding site may cause errors in analysis. The aim of this study was to determine the prevalence of the variants in a Danish cohort examined for lactose intolerance as well as to improve the real-time PCR analysis for detection of the different variants. METHODS: We genotyped 3395 routine samples using real-time PCR for the -13910C > T-variant. All consecutive samples identified as -13910CC were sequenced using Sanger Sequencing. Using the SDS software we examined various quality value settings to improve on the genetic analysis. RESULTS: Using real-time PCR resulted in 100% successful genotyping of the -13910C > T variant. By using a quality value of 99% and sequencing the undetermined samples we improved the ability of the assay to identify variants other than -13910C > T. This resulted in a reduction of the diagnostic error rate by a factor of 2.4 while increasing the expenses only 3%. CONCLUSIONS: We conclude that using a quality value of 99% in the SDS software significantly improves the diagnostic efficiency of the real-time PCR assay for detecting variants associated to lactase persistence.

Consumption of milk and dairy products: Facts and figures.

OBJECTIVES: Consumption of milk has been declining sharply in recent decades, particularly in developed countries. One of the reasons for this decline is the diagnosis or perception of lactose intolerance. The aim of this study was to investigate average consumption of milk and dairy products in the Campania region of Italy, one of the main producers of dairy products in the country. METHODS: Individuals aged 18 to 75 y and living in Campania were invited to answer an online questionnaire regarding their average consumption of milk and dairy products. The questionnaire was posted on the public access hospital site, as well as on several Facebook pages of friends and hospital personnel. RESULTS: The study found that 22.2% (260 of 1173) of responders from Campania do not drink milk, and 18.1% (213 of 1173) drink lactose-free milk, mainly because of gastrointestinal symptoms. The vast majority of the sample population chose to avoid consuming milk without undergoing the breath test for lactose intolerance or consulting a doctor. Women and underweight people drink more lactose-free milk than milk containing lactose. The population sample does not avoid dairy products; rather, they seem to be consumed quite frequently. CONCLUSION: The data support the need for mandatory implementation of a nutritional campaign to increase understanding regarding, for example, unnecessary avoidance of milk and excessive consumption of cheese.

Milk hypersensitivities: where is the grey line regarding their dietary management?

The proportion of people suffering or reporting to have a hypersensitivity caused by cow's milk consumption is increasing, and even health professionals often face difficulties into elaborating properly with a milk reaction due to misdiagnosis. The scope of this review is to present literature data that lead into putting the border line between cow's milk allergy and cow's milk intolerance, mainly focusing on how the different pathophysiology leads to their different dietary diagnosis and management.

Food avoidance in athletes: FODMAP foods on the list.

We surveyed 910 athletes to assess behaviours towards self-selected food/ingredient avoidance to minimize gastrointestinal distress. Fifty-five percent eliminated at least 1 high fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (FODMAP) food/category, with up to 82.6% reporting symptom improvement. In athletes indicating that high FODMAP foods trigger gastrointestinal symptoms, lactose (86.5%) was most frequently eliminated, followed by galactooligosaccharides (23.9%), fructose (23.0%), fructans (6.2%), and polyols (5.4%). Athletes avoid predominantly lactose and to a lesser extent other high FODMAP foods to reduce gastrointestinal distress.

Prevalence and Symptom Correlation of Lactose Intolerance in the North East Part of Bangladesh.

This study was designed to see the prevalence of lactose intolerance and symptom correlation following oral lactose challenge in healthy volunteers in the north east part of Bangladesh. Symptoms of abdominal pain, nausea, borborygmi, flatulence, diarrhea and others were noted for 24 hours and blood glucose was estimated at 0 hour and 30 minutes after 50 gm oral lactose load to healthy volunteers. Failure to rise blood glucose level ≥1.1 mmol/l at 30 minutes after lactose intake from fasting level was taken as lactose malabsorption (LM) i.e., lactose intolerance. Sensitivity and specificity of different symptoms were then found out. A total of 171 volunteers (male 123, female 48) with a mean age 34.08 years participated in this study. Lactose intolerance was found among 82.5% (n=141, M=100, F=41) subjects. Symptoms mostly experience by the lactose malabsorbers were diarrhea 93(66.0%), borborygmi 80(56.7%), abdominal pain 31(22.0%) and flatulence 32(22.7%). LM prevalence was found to increase with increasing number of symptoms up to 3 symptoms. A week positive correlation (r=0.205, P=0.007) was found between the number of symptoms and proportion of subjects having positive lactose tolerance test. Lactose intolerance among healthy adults of North East part of our country is as common as in other Asian countries including China and Malaysia. But LM is higher than that of Europeans and south Indians. Diarrhea and borborygmi were mostly associated with LM.

From 'lactose intolerance' to 'lactose nutrition'.

The concept of lactose intolerance has become embedded in Western medicine and developing economy medicine. It is based on evidence that intestinal lactase activity persists into later childhood and throughout life in only a minority of the world's population, notably northern European-derived populations. These people have the T single nucleotide polymorphism (SNP) of the rs49882359 allele (C/T), also known as C/T-13910, the MCM6 gene which positively influences the lactase LCT gene. Other lactase persistent (LP) populations are found in Africa and the Middle East with different genetic variants. These SNPs represent co-evolution with dairying since the agricultural revolution and nutrient-dependent ecological adaptation. That said, gastrointestinal symptoms considered due to small intestinal lactose malabsorption are poorly correlated with lactase non-persistence (LNP), the situation for most people. With LNP, colonic microbiome lactase enables lactose fermentation to occur so that none is found in faeces. Whether the short chain fatty acids (SCFAs) and gases (hydrogen, carbon dioxide and methane) produced cause symptoms is dose-dependent. Up to 25 g of lactose at any one time can usually be consumed by a LNP person, but its food and meal pattern context, the microbiomic characteristics, age and other factors may alter tolerance. Thus, the notion that lactose intolerance is a disorder or disease of LNP people is misplaced and has been one of cultural perspective. What actually matters is whether a particular dairy product as normally consumed give rise to symptoms. It is, therefore, proposed that lactose tolerance tests be replaced with dairy food tolerance tests.

Lactose intolerance in Indonesian children.

"Lactose intolerance (LI)" is considered a common problem in Asians, and in many parts of the world. Its prevalence and age of manifestation varies between by Asian country, for possible genetic or cultural reasons. Studies in Indonesian children 3-15 years old (y) are available within the past two decades, using a pure lactose tolerance test. The prevalences of lactose malabsorption (LM) in pre-elementary (3-5 y), elementary (6-11 y), and junior high (12-14 y) school-children were 21.3%, 57.8%, and 73%, respectively. An increasing trend for LM prevalence was seen within the pre-elementary group, from 9.1% at 3 y to 28.6% at 5 y. The most frequent symptoms of LI in junior high school (JHS) group were abdominal pain (64.1%), abdominal distention (22.6%), nausea (15.1%), flatulence (5.7%), and diarrhea (1.9%), mostly within one hour of lactose ingestion. In children with regular and irregular milk drinking, LM occurred in 81.2% and 69.6%; LI was found in 56.2% and 52.1%, respectively. Most JHS children with dairy-associated recurrent abdominal pain (RAP) symptoms proved to be malabsorbers. Dairy products most related to RAP were milk and yogurt. LI was found in 81% of RAP children with abdominal pain most frequently, followed by nausea, bloating, diarrhea, borborygmi, and flatulence. Symp-tom onset occurred 30 minutes after lactose ingestion, especially nausea, bloating, and abdominal pain. In RAP children LI symptoms mostly found in breath hydrogen concentration>20 ppm. More LI symptoms were found in lactose malabsorbers, but symptoms were mild and generally disappeared in 7 hours, and in most by 15 hours.

Additional Value of CH4 Measurement in a Combined (13)C/H2 Lactose Malabsorption Breath Test: A Retrospective Analysis.

The lactose hydrogen breath test is a commonly used, non-invasive method for the detection of lactose malabsorption and is based on an abnormal increase in breath hydrogen (H2) excretion after an oral dose of lactose. We use a combined (13)C/H2 lactose breath test that measures breath (13)CO2 as a measure of lactose digestion in addition to H2 and that has a better sensitivity and specificity than the standard test. The present retrospective study evaluated the results of 1051 (13)C/H2 lactose breath tests to assess the impact on the diagnostic accuracy of measuring breath CH4 in addition to H2 and (13)CO2. Based on the (13)C/H2 breath test, 314 patients were diagnosed with lactase deficiency, 138 with lactose malabsorption or small bowel bacterial overgrowth (SIBO), and 599 with normal lactose digestion. Additional measurement of CH4 further improved the accuracy of the test as 16% subjects with normal lactose digestion and no H2-excretion were found to excrete CH4. These subjects should have been classified as subjects with lactose malabsorption or SIBO. In conclusion, measuring CH4-concentrations has an added value to the (13)C/H2 breath test to identify methanogenic subjects with lactose malabsorption or SIBO.

Contribution of Colonic Fermentation and Fecal Water Toxicity to the Pathophysiology of Lactose-Intolerance.

Whether or not abdominal symptoms occur in subjects with small intestinal lactose malabsorption might depend on differences in colonic fermentation. To evaluate this hypothesis, we collected fecal samples from subjects with lactose malabsorption with abdominal complaints (LM-IT, n = 11) and without abdominal complaints (LM-T, n = 8) and subjects with normal lactose digestion (NLD, n = 15). Lactose malabsorption was diagnosed using a (13)C-lactose breath test. Colonic fermentation was characterized in fecal samples at baseline and after incubation with lactose for 3 h, 6 h and 24 h through a metabolomics approach using gas chromatography-mass spectrometry (GC-MS). Fecal water cytotoxicity was analyzed using a colorimetric assay. Fecal water cytotoxicity was not different between the three groups (Kruskall-Wallis p = 0.164). Cluster analysis of the metabolite patterns revealed separate clusters for NLD, LM-T and LM-IT samples at baseline and after 24 h incubation with lactose. Levels of 5-methyl-2-furancarboxaldehyde were significantly higher in LM-IT and LM-T compared to NLD whereas those of an unidentified aldehyde were significantly higher in LM-IT compared to LM-T and NLD. Incubation with lactose increased short chain fatty acid (SCFA) concentrations more in LM-IT and LM-T compared to NLD. In conclusion, fermentation patterns were clearly different in NLD, LM-IT and LM-T, but not related to differences in fecal water cytotoxicity.

Milk Intolerance, Beta-Casein and Lactose.

True lactose intolerance (symptoms stemming from lactose malabsorption) is less common than is widely perceived, and should be viewed as just one potential cause of cows' milk intolerance. There is increasing evidence that A1 beta-casein, a protein produced by a major proportion of European-origin cattle but not purebred Asian or African cattle, is also associated with cows' milk intolerance. In humans, digestion of bovine A1 beta-casein, but not the alternative A2 beta-casein, releases beta-casomorphin-7, which activates µ-opioid receptors expressed throughout the gastrointestinal tract and body. Studies in rodents show that milk containing A1 beta-casein significantly increases gastrointestinal transit time, production of dipeptidyl peptidase-4 and the inflammatory marker myeloperoxidase compared with milk containing A2 beta-casein. Co-administration of the opioid receptor antagonist naloxone blocks the myeloperoxidase and gastrointestinal motility effects, indicating opioid signaling pathway involvement. In humans, a double-blind, randomized cross-over study showed that participants consuming A1 beta-casein type cows' milk experienced statistically significantly higher Bristol stool values compared with those receiving A2 beta-casein milk. Additionally, a statistically significant positive association between abdominal pain and stool consistency was observed when participants consumed the A1 but not the A2 diet. Further studies of the role of A1 beta-casein in milk intolerance are needed.

[THERAPY OF TRANSIENT LACTASE INSUFFICIENCY OF CHILDREN IN PECTORAL AGE].

Thus, we have discovered that the children of the first half-year of life have different degrees of severity of transient lactase insufficiency basing on the results of hydrogen respiratory test. It was set that the starting dose of enzyme lactase must depend on the degree of severity of displays of transient lactase insufficiency, taking into account the indexes of hydrogen respiratory test.

Small intestinal bacterial overgrowth as an uncommon cause of false positive lactose hydrogen breath test among patients with diarrhea-predominant irritable bowel syndrome in Asia.

BACKGROUND AND AIM: It has been reported that small intestinal bacterial overgrowth (SIBO) may lead to false positive diagnoses of lactose malabsorption (LM) in irritable bowel syndrome patients. The aim of this study was to determine the influence of SIBO on lactose hydrogen breath test (HBT) results in these patients. METHODS: Diarrhea-predominant irritable bowel syndrome patients with abnormal lactose HBTs ingested a test meal containing (99m) Tc and lactose. The location of the test meal and the breath levels of hydrogen were recorded simultaneously by scintigraphic scanning and lactose HBT, respectively. The increase in hydrogen concentration was not considered to be caused by SIBO if ≥ 10% of (99m) Tc accumulated in the cecal region at the time or before of abnormal lactose HBT. RESULTS: LM was present in 84% (31/37) of irritable bowel syndrome patients. Twenty of these patients agreed to measurement of oro-cecal transit time. Only three patients (15%) with abnormal lactose HBT might have had SIBO. The median oro-cecal transit time between LM and lactose intolerance patients were 75 min and 45 min, respectively (Z=2.545, P=0.011). CONCLUSIONS: Most of irritable bowel syndrome patients with an abnormal lactose HBT had LM. SIBO had little impact on the interpretation of lactose HBTs. The patients with lactose intolerance had faster small intestinal transit than LM patients.

An insight into the gastrointestinal component of fibromyalgia: clinical manifestations and potential underlying mechanisms.

Fibromyalgia syndrome is characterized by chronic generalized pain accompanied by a broad symptomatologic spectrum. Besides chronic fatigue, sleep disturbances, headaches and cognitive dysfunction that are extensively described in the literature, a considerable proportion of patients with fibromyalgia experience gastrointestinal symptoms that are commonly overlooked in the studies that are not specifically dedicated to evaluate these manifestations. Nevertheless, various attempts were undertaken to explore the gastrointestinal dimension of fibromyalgia. Several studies have demonstrated an elevated comorbidity of irritable bowel syndrome (IBS) among patients with fibromyalgia. Other studies have investigated the frequency of presentation of gastrointestinal symptoms in fibromyalgia in a nonspecific approach describing several gastrointestinal complaints frequently reported by these patients such as abdominal pain, dyspepsia and bowel changes, among others. Several underlying mechanisms that require further investigation could serve as potential explanatory hypotheses for the appearance of such manifestations. These include sensitivity to dietary constituents such as gluten, lactose or FODMAPs or alterations in the brain-gut axis as a result of small intestinal bacterial overgrowth or subclinical enteric infections such as giardiasis. The gastrointestinal component of fibromyalgia constitutes a relevant element of the multidisciplinary pathophysiologic mechanisms underlying fibromyalgia that need to be unveiled, as this would contribute to the adequate designation of relevant treatment alternatives corresponding to these manifestations.