The toll-like receptor 4 Asp299Gly variant: no influence on LPS responsiveness or susceptibility to pulmonary tuberculosis in The Gambia.

SETTING: Tuberculosis (TB) remains a major cause of morbidity and mortality worldwide. Studies in a murine model of pulmonary TB have identified a role for Toll-like receptor 4 (TLR4) in the development of chronic lung infection with Mycobacterium tuberculosis. The Asp299Gly polymorphism in the human TLR4 gene is associated with in vivo hyporesponsiveness to lipopolysaccharide (LPS) in Caucasians. OBJECTIVE: To determine whether TLR4 Asp299Gly influences LPS responses or susceptibility to pulmonary TB in humans in a Gambian population sample. DESIGN: We compared whole blood monokine responses to LPS in 245 healthy blood donors stratified by TLR4 Asp299Gly genotype to assess whether this polymorphism was functional in this population. A case-control study of 640 subjects was conducted to investigate whether TLR4 Asp299Gly was associated with TB. RESULTS: LPS-induced tumour necrosis factor, interleukin-1 beta and interleukin-10 production was not influenced by TLR4 Asp299Gly genotype. There was no association between TLR4 Asp299Gly and TB. CONCLUSION: Our data suggest that TLR4 Asp299Gly has no influence on monocyte LPS responses or susceptibility to TB in Gambians and could be an ancient neutral polymorphism.

Are all hospital admissions for acute gastroenteritis necessary?

Consecutive hospital admissions of 300 children under 2 years old with gastroenteritis were studied to establish the reasons for the request for admission, treatment practices before admission, and the severity of illness. Symptoms were mostly mild. Inappropriate treatment at home was unusual but 87 (29%) had received none before being admitted. In 66 cases (22%) more than one reason was given for referral and 61 (20%) were referred in part for non-medical reasons. Hospital stay was often longer than necessitated by the severity of the illness alone. Redeployment of resources to fund assessment centres on the paediatric ward and more community nursing teams would allow many more of these children to be nursed safely at home.

Experience of a specialist service for advice on childhood immunisation.

We report a 3 year experience of a specialist clinic set up to advise on childhood immunisation. In all, 20 children were referred for advice on their suitability for measles, mumps, rubella (MMR) immunisation and 93 for advice about pertussis immunisation. All of the former and 78 (84%) of the latter were advised that there were no contraindications to their immunisation. The 20 children given MMR vaccine and 55 (71%) of the 78 who were immunised against pertussis had no adverse effects. The clinic serves a small but important group of children who, because of parental or health care professional uncertainty, may not receive protection against potentially damaging or fatal infections.

Incidence of rotavirus in artificially reared pigs and some effects of diarrhoea on the physiology and histology of the gastrointestinal tract.

Pigs reared on a milk substitute from two days old often developed diarrhoea, but suckled littermates remained healthy. Only in a few pigs was diarrhoea associated with the presence of rotavirus. Rotavirus was also present in some healthy pigs, and was associated with a reduction in villus length. Pigs with diarrhoea usually had an increased amount of digesta in the stomach and a reduction in lactase activity in the small intestine but villus length was unchanged. There was no evidence of lactose malabsorption.

The aetiology of diarrhoea in pigs weaned at two days of age.

When pigs are weaned at two days of age large numbers of Excherichia coli appear in the anterior gut and the incidence of diarrhoea rises. The two phenomena do not appear to be directly related because the strains of E coli isolated are not serotypes previously found to be associated with neonatal pig scouring. Representative strains of the non-enteropathogenic serotypes did not produce enterotoxin and did not adhere to small intestine brush borders. Moreover when antibiotics were fed to eliminate E coli from the gut, the pigs still scoured. Rotavirus was detected in the gut contents and gut epithelium of scouring pigs and a bacteria-free filtrate of gut contents produced diarrhoea when administered to germ-free pigs. It is suggested that rotavirus may be one of the causes of the scouring seen shortly after weaning pigs at two days of age.

Genetics of susceptibility to tuberculosis in humans.

There is substantial epidemiological evidence that host genetic factors in part determine susceptibility to mycobacteria, and many approaches have been applied to identify the specific genes involved. These include the study of single genes in 'knockout' mouse models and rare human families in which increased susceptibility to mycobacterial infection segregates as a single gene defect. Several genes have now been studied in many different populations. This review gives an overview of the progress made in the field of genetic susceptibility to tuberculosis and highlights more generally some of the challenges involved in the identification of complex disease genes.

Hepatobiliary infections due to non-capsulated Haemophilus influenzae.

We present two cases of non-capsulated Haemophilus influenzae hepatobiliary infection and review the literature. Such cases are rare, and prior to routine immunization against H. influenzae serotype b invasive Haemophilus disease was largely caused by capsulated strains. The epidemiology of invasive Haemophilus infections has changed and the number of cases of intra-abdominal and hepatobiliary infection may be underestimated due to current microbiological processing practices.

Capacity-building in human genetics for developing countries: initiatives and perspectives in sub-Saharan Africa.

BACKGROUND: Stakeholders who are committed to bridge the gap in genetics services need to be aware of current initiatives in sub-Saharan Africa. METHODS: We reviewed selected experiences from African geneticists that led to specific recommendations. RESULTS: The initiation of prenatal diagnosis of sickle cell anaemia founded the first medical genetic service in Cameroon. There remains a need for international collaborative effort to overcome the lack of human, technical and financial resources around the practice of medical genetics in Africa. The African Society of Human Genetics, Wellcome Trust and NIH have recently proposed a model on how to fully engage Africa in genomics. It includes a 'Health and disease' phase I: use of the case-control design to study genetic and epidemiological determinants of 7 important diseases in Africa, and a 'Genetic variation' phase II: comprehensive documentation of genetic variations in 100 carefully selected ethnic groups across Africa. The strategy would require the development of: (1) clinical phenotyping centres, (2) molecular phenotyping centres, (3) genotyping and sequencing capability, (4) data centres, and (5) a bio-repository in Africa. CONCLUSIONS: Governments and international health agencies need to recognise that genetics is important to the global medical community. The initiatives of African geneticists need advocacy and encouragement from the international community.

Global distribution of a novel trinucleotide microsatellite polymorphism (ATA)n in intron 8 of the SLC11A1 gene and susceptibility to pulmonary tuberculosis.

We identified a novel trinucleotide (ATA)n repeat polymorphism in intron 8 of SLC11AI, a candidate gene for susceptibility to tuberculosis (TB) infection. We characterized the frequency of this polymorphism in 485 individuals originating from eight globally diverse human populations and compared the distribution of (ATA)n alleles in 146 adults and in 80 cord blood samples from newborns in the Gambian population. Lastly, we tested for association of this microsatellite with pulmonary TB in 318 TB cases and 146 controls in the Gambian population. We found no significant difference in frequency or distribution of alleles in adult and cord blood samples, and we found no significant association between this marker and pulmonary TB.

Influence of genetic and environmental factors on the immunogenicity of Hib vaccine in Gambian twins.

The differences in incidence rates of Haemophilus influenzae type b disease and the variation in Hib conjugate vaccine efficacy achieved among different ethnic groups suggest genetic influences on the immune response to Hib vaccine. The serum anti-PRP antibody concentration of 43 monozygotic (MZ) and 147 dizygotic (DZ) twin pairs in the Gambia was measured using a standardised Hib ELISA. Intrapair correlations for MZ and DZ twin pairs were compared and heritability in antibody responses to Hib conjugate vaccine was estimated to be 51% (95% CI: 32-66%), indicating a significant genetic contribution in the response. We conclude that genetic factors may be involved in the variation in immune response to Hib vaccine observed in different populations and may contribute to cases of vaccine failure.

Molecular diagnosis of Fusobacterium necrophorum infection (Lemierre's syndrome).

Presented here is the case of a 27-year-old male with atypical features of Lemierre's syndrome in which a definitive diagnosis was achieved using molecular methods. While routine investigations, including bacterial cultures, were unhelpful, two real-time PCR assays demonstrated Fusobacterium necrophorum-specific DNA in aspirates from brain and renal abscesses. This is the first report demonstrating that a laboratory diagnosis can be made using molecular methods in suspected cases of Lemierre's syndrome. Use of these methods can thus resolve diagnostic confusion, prevent unnecessary investigation, and direct specific antimicrobial treatment.

Artificial rearing of pigs. 11. Effect of replacement of dried skim-milk by an isolated soya-bean protein on the performance of the pigs and digestion of protein.

1. Pigs (twenty-one/diet) were weaned at 2 d of age and given liquid diets (200 g dry matter/l) at hourly intervals during a 26 d experiment. The pigs were fed on a scale based on live weight. The diets contained (g/kg DM): dried skim-milk 730 (diet A), dried whey 508.5, isolated soya-bean protein 218, DL-methionine 3.5 (diet S), and soya-bean oil 270 (diets A and S). Diet T contained equal proportions of diets A and S. Soya-bean supplied 0, 370 and 740 g crude protein (nitrogen x 6.25) kg total crude protein in diets A, T and S respectively. 2. Performance was similar for both diets A and T (P > 0.05). Pigs given diet S scoured severely, and fourteen died. The survivors grew very poorly. Nitrogen retention (g/d per kg live weight) was greater for diet A compared with diet T (P < 0.01), and decreased with age (P < 0.001). 3. Protein digestion was examined in the pigs killed at 28 d of age. The amount of soya-bean protein in the diet did not affect the amount of digesta in the stomach, but soya-bean protein decreased the pH, DM and total N content of the digesta (P < 0.01), and increased, though not significantly (P > 0.05), pepsin activity in the digesta and stomach tissue. Acid secretion into the stomach appeared to be enhanced in pigs given a diet containing soya-bean protein. 4. Amounts of trypsin, chymotrypsin, total N and proportion of non-protein-N in the digesta from the small intestine were similar for both diets A and T. The amounts for both diets were greater in the distal compared with the proximal region of the small intestine (P < 0.05). Chymotrypsin activity in the pancreas was reduced (P < 0.05) in pigs given diet T, although this reduction did not seem to impair digestion in 28-d-old pigs. Trypsin activity in the pancreas was similar for both diets A and T. 5. It seems likely that the neonatal pig does not have the digestive capacity to tolerate the large daily intakes of soya-bean protein when dried skim-milk was totally replaced in the diet (diet S). When half the dried skim-milk was replaced, protein digestion was not impaired in 28-d-old pigs.

Growth, digestion, and protein metabolism in neonatal pigs given diets containing whey as the predominant or only source of milk protein.

Milk substitutes containing ratios of casein/whey protein ranging from 80:20 to 0:100 were given to neonatal pigs. A ratio of 60:40 gave maximum growth rate, efficiency of feed utilization and nitrogen retention, and the lowest concentration of urea in blood plasma. This ratio is close to that in sow's milk, suggesting the hypothesis that in each species the milk may be adapted to the protein requirements of their young; by analogy, a casein/whey protein ratio of 20:80 in humanized milk formula might lead to more efficient protein utilization by the infant. In general, changes in the proportion of casein and whey proteins in the diet produced similar effects on the free amino acids in blood plasma as were found in clinical studies reported in the literature, providing further evidence of similarities in the protein metabolism of infants and neonatal pigs. The amount of nitrogen in the digesta remaining in the stomach at 1 h after a meal indicated that whey proteins empty from the stomach more rapidly than casein.

Vitamin B12 absorption in the neonatal piglet. 2. Resistance of the vitamin B12-binding protein in sows' milk to proteolysis in vivo.

It has been postulated that the vitamin B12-binding protein in the sows' milk may facilitate the intestinal absorption of vitamin B12 in the piglet. This implies that the binder is not rapidly and completely degraded by gut proteases, and the present experiments were devised to test this assumption. Piglets aged 7 and 28 d were given a test feed of sows' milk whey both with and without cyano[3H]cobalamin to saturate the binder, and with 14C-labelled polyethylene glycol (PEG) as a marker. Control piglets were given a milk substitute containing no vitamin B12-binder. At 80 min after the test meal the piglets were killed and the contents of the stomach and small intestine were removed for analysis. Recovery of [14C]PEG, mainly from the middle and distal portions of the small intestine, ranged from 49 to 78%. Apparent loss of vitamin B12-binder from the intestine was calculated from the ratio, PEG:binder. In both 7- and 28-d-old piglets, the content of saturated binder in the middle and lower small intestine was about 20% depleted relative to that in the test fed. With the unsaturated binder the loss was more variable and generally greater, at about 50%. At slaughter the stomach was nearly empty and the high unsaturated binding capacity of the contents was attributed to endogenous binders. However, in the middle and lower small intestine, the binding capacity was derived mainly from sows' milk given with the test meal. No free cyano[3H]cobalamin was found in the intestinal contents of piglets given sows' milk to which enough cyano[3H]cobalamin had been added to saturate the binder. There was no change in the molecular weight of the vitamin B12-binder during its passage down the intestine, as judged by its behaviour on filtration in Sephadex gel G-150. The results indicate that a high proportion of saturated binder, and a smaller proportion of unsaturated binder, survived unchanged in the intestine of piglets at both ages.

Artificial rearing of pigs. 12. Effect of replacement of dried skim-milk by either a soya-protein isolate or concentrate on the performance of the pigs and digestion of protein.

1. Pigs (sixteen/diet) were weaned at 2 d of age and given liquid diets (200 g dry matter/l) during a 26 d experiment. The pigs were fed on a scale based on live weight. Dried skim-milk was the only source of protein in diet U and was partially or totally replaced by a soya-bean isolate (diet B) or a concentrate (diets C and D). Soya-bean protein provided 500, 700 or 350 g/kg total crude protein (nitrogen x 6.25) in diets B, C and D respectively. 2. Performance was similar for diets B and D, but poorer than that of pigs given diet U. The apparent digestibility and retention of N of these diets was similar. Pigs given diet C scoured severely and twelve died. 3. Protein digestion was studied in pigs given diets U, B and D, killed at 28 d of age, at the termination of the feeding experiment. The dry matter content and proportion of N in the digesta in the stomach were reduced in pigs given soya-bean protein. Pepsin concentrations in digesta and stomach tissue were unchanged. 4. The concentrations of trypsin and chymotrypsin in the pancreas were greater in pigs given the soya-bean protein concentrate compared with milk protein, but only the increase in trypsin was significant (P less than 0.01). Digesta from the small intestine of pigs given the soya-bean-protein isolate contained less chymotrypsin (P less than 0.05). There were no differences in the proportion of non-protein-N in the total N in the digesta, suggesting that proteolysis of the milk and soya-bean proteins were equally by 28 d of age.

Evaluation of the neonatal pig as a model for infant nutrition: effects of different proportions of casein and whey protein in milk on nitrogen metabolism and composition of digesta in the stomach.

Neonatal pigs were evaluated as a model for protein metabolism of infants. Milks were given in which the protein (15 or 30 g/liter) was supplied by casein and whey in the proportions of 80:20 or 40:60. Severe or moderate restriction of feed intake was also compared. Whey-predominant milks reduced the concentration of urea N in blood plasma and the proportion of urea N in urinary total N. These effects are consistent with most previously reported data from infants, particularly those of low birth weight. The efficiency of N retention was improved at the higher concentration of dietary protein. Whey-predominant milks also reduced the amount of N retained in the stomach. The magnitude of some of these effects was influenced by the level of feed intake. Total N retention reflected protein intake but was not affected by protein source. N retention determined by a balance procedure exceeded that calculated from carcass analysis, but the effect of protein intake was similar from either procedure.

Gastric proteases in the human infant.

The electrophoretic mobilities of proteases present in gastric juice taken within 10 h of birth from 5 healthy, premature infants were compared with calf chymosin, pig pepsin A and human adult gastric juice. The juice from 2 infants contained predominantly a chymosin-like enzyme, another had almost exclusively pepsins similar to those of the adult juice, while the other two contained a mixture of both. The pepsins consisted of two elements, probably pepsin A (EC 3.4.23.1), and pepsin C (EC 3.4.23.3). Single radial immunodiffusion gave a definite reaction to calf anti-chymosin serum in five samples taken from a further 17 infants. These results indicate that some human infants secrete chymosin. The reaction in the immunodiffusion assay indicated a much lower enzyme activity than that implied from electrophoretic separations. It is suggested that species differences resulted in poor cross-reactivity of the antiserum.

Variation in the tumor necrosis factor-alpha gene promoter region may be associated with death from meningococcal disease.

Tumor necrosis factor-alpha (TNF-alpha) plays a central role in the pathophysiology of sepsis. Levels of TNF-alpha are directly correlated with severity in meningococcal disease (MD). A polymorphism in the promoter region of the TNF-alpha gene is associated with differences in the secretion of TNF-alpha. The TNF2 allele is associated with higher constitutive and inducible levels of TNF-alpha secretion than is the TNF1 allele. To investigate whether possession of the TNF2 allele is associated with severity in MD, the frequency of TNF1 and TNF2 alleles in 98 children with MD was compared. There were more deaths among children who had the TNF2 allele (P = .03; relative risk [RR], 2.5; 95% confidence interval [CI], 1.1-5.7) than in those who did not. There was also an increased risk of severe disease in children with the TNF2 allele (P = .02; RR, 1.6; 95% CI, 1.1-2.3). Possession of the TNF2 allele predisposes to a worse outcome in children with meningococcal infection.

Artificial rearing of pigs. The effect of different levels of fat, protein and methionine in a milk-substitute diet containing skim milk and soya-bean oil.

1. Protein and energy requirements of pigs weaned at 2 d of age were studied. In Expt 1, three diets with crude protein (CP; nitrogen x 6.38) and fat contents (g/kg dry matter (DM)) of 268, 260 (diet A); 216, 415 (diet B) and 264, 405 (diet C) were compared, with and without a supplement of 2 g DL-methionine/kg. Diets A and B contained skim milk and soya-bean oil only, in different proportions. Casein was added to diet C to equalize the CP content to that of diet A. In Expt 2, diets containing 290 or 315 g CP/kg DM were used with either 310 or 405 g fat/kg. The diets contained different proportions of skim milk, soya-bean oil and casein. The same diets as used in Expt 2 were used in Expt 3 but fed to a scale based on live weight which was 25% less than that used in Expts 1 and 2. 2. Methionine supplementation had no significant effect (P greater than 0.05) on the growth rate or feed: gain ratio of pigs from 2 to 28 d of age. The growth rate of the pigs from 2 to 28 d of age was significantly (P less than 0.05) reduced when they were given the higher level of either CP or fat, with the exception that the effect of fat was not significant (P greater than 0.05) in Expt 3. The reduction in growth rate was associated with a decrease in feed intake. Feed: gain ratios were unaffected (P greater than 0.05) by the level of either CP or fat in the diet. 3. The higher protein level did increase the proportion of CP in the carcass DM (Expt 2 P less than 0.01, Expt 3 P greater than 0.05) and gave an indication, which was not significant (P greater than 0.05), of improved N retention (g/d per kg live weight). The higher dietary fat level increased the proportion of lipid in the carcass DM, and this effect was significant (P less than 0.05) in both Expts 2 and 3. 4. Diets containing additional casein were not fully consumed at the level of feeding used in Expt 2. This effect may be associated with the larger amounts of digesta in the stomachs of these pigs. Some differences in the composition of the digesta in the stomach were observed, but these did not affect the performance of the pigs. 5. It seems likely that either growth rate or feed: gain ratio (g DM consumed/g live-weight gain) can be improved by protein or energy levels exceeding those in a diet containing (g/kg DM) 730 skim milk (265-290 CP) and 310 fat. In this diet, containing (g/kg DM) 6-4 methionine and 3-5 cystine, sulphur amino acid requirements were satisfied.