Paediatrics and Child Health

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Nephrotic syndrome in African and Indian children in South Africa.
(1981) Adhikari, Miriam.
There are comprehensive accounts of the nephrotic syndrome in childhood in temperate countries. Many of the important features of this disease have been known for close on to two decades. The causal link between malaria and nephrosis in tropical Africa has also been recognised and documented for a similar length of time. Very little was known of the nephrotic syndrome in the sub-tropical zones of Africa where malaria is not endemic. Anecdotal evidence in South Africa suggested that African children with this disease appeared to have steroid resistant nephrosis and a more protracted clinical course than expected from prevailing accounts in the literature and that Indian South African children generally responded to steroids. This thesis is the result of detailed investigations in to this disease in African and Indian children in Durban, South Africa. 2. Preliminary Study A preliminary study was undertaken in which 53 (12 African and 41 Indian) children with the nephrotic syndrome defined by clinical and biochemical criteria 1 ii were studied. Renal biopsies were not available on these patients. The results revealed that two thirds of the African children were over 5 years of age and 50% were males. Of the Indian children 50% were under 5 years of age and 50% were males. Nine African children were treated with steroids and 8 did not respond whereas 31 of the 39 Indian children treated clearly responded to steroid therapy. In addition 5 Indian patients were treated with cyclophosphamide and 3 responded. On follow-up 7 of the African children had persistent proteinuria, 2 experienced remissions and 3 were lost to follow-up. All the Indian patients experienced remissions. The differences between the 2 groups of nephrotic patients were quite striking and therefore a more detailed prospective study of this problem was undertaken. 3. Prospective Study of Primary Nephrotic Syndrome One hundred and seventy children of whom 104 were African and 66 Indian with primary nephrotic syndrome were studied. In both racial groups the male sex dominated, Indian children tended to present iii at a younger age group whereas African children presented at two peak ages, 5 years and between 5 - 10 years. 3.1 Histological Differences The histological types found on light microscopic examination of renal tissue were distinctly different between the African and the Indian children. The majority (85.6%) of the African children had 'obvious' glomerular lesions, the commonest being extramembranous nephropathy (29.8%). Although the proliferative group was the single largest group (40%) none of the subgroups exceeded the extramembranous type in their number. Minimal change accounted for only 14.4% of the African children with nephrotic syndrome. The majority of Indian children (72.7%) had minimal change on light microscopy, 9.1% focal glomerulosclerosis and 12.1% had proliferative changes. 3.2 Immunofluorescence Immunofluorescent studies also indicated differences between the two groups of patients. Generally, heavier deposits of immunoglobulins iv and complement components were identified on renal biopsy specimens of African children. This occurred even in MCNS where most African children had heavy IgG, light IgM, IgA and complement components whereas only a few of the Indian children had light IgM deposits. Similar differences were observed in diffuse mesangial proliferative glomerulonephritis and focal glomerulosclerosis where the numbers of patients were comparable. 3.3 Presenting Features Clinical features at presentation in the two groups were different, as expected from the nature of the histological findings in each group. In the African children (all histological groups) haematuria occurred in 35.5%, hypertension 16.3% and renal failure in 2.9%. The clinical features in the Indian children were not too different from MCNS elsewhere. Haematuria occurred in a small percentage (3%) of MCNS but was more frequent (10.7%) in other groups. Hypertension and renal failure occurred infrequently in histological categories other than MCNS where they did not occur at all. 3.4 Course and Outcome In view of the above it was not unexpected to find that the clinical course and outcome in the two groups were quite dissimilar. African patients in certain of the histological groups fared reasonably well, but none of the groups had the excellent prognosis of Indian MCNS. 3.4.1 Minimal Change One third of the African MCNS patients remitted and this was unrelated to steroids. The remainder who were followed for a reasonable duration of time remained proteinuric. None developed signs of serious renal impairment (azotaemia, hypertension). Indian MCNS experienced an excellent prognosis with 97.8% achieving remission and 81.6% being steroid sensitive. One third of these patients had a single episode of nephrosis while frequent relapses occurred in 28.2%. 3.4.2 Extramembranous Nephropathy Patients with extramembranous nephropathy, the largest group in the African patients, experienced hypertension more often (20%) vi and remission less often (30%) than do children in temperate climates. The clinical presentation, course and outcome in the majority of these patients were similar to adults with extramembranous nephropathy. 3.4.3 Proliferative Glomerulonephritis The patients in the proliferative group had a variable outcome depending on the subgroup to which they belonged. In diffuse mesangial proliferation, African patients had a higher incidence of hypertension and fewer remissions and fared less well than Indian patients. The diffuse endocapillary glomerulonephritis, membranoproliferative and focal proliferative nephritis groups of patients suffered severe disease with a failure to remit and progression to death. In the diffuse exudative group, remissions occurred or proteinuria persisted but severe relapse and death did not occur. The worst prognosis was in the focal proliferative group with the highest incidence of persistent relapse. 3.4.4 Focal Glomerular Sclerosis Focal glomerular sclerosis was an unusual vii histological diagnosis in the African child (3.9%) with a poorer prognosis (persistent proteinuria or death) when compared to Indian children in whom one third remitted and the rest had persistent proteinuria. 3.4.5 Tropical Nephropathies It is difficult to comment on the course of the tropical nephropathy (not related to malaria) and tropical extramembranous groups as the numbers are small. However, in tropical extramembranous, none remitted (all African children) and in tropical nephropathy one Indian child remitted but one of 2 African children died and the other had persistent proteinuria. 3.5 Response to Therapy Perhaps the most important practical aspect of the nephrotic syndrome in the African child was the response to steroid therapy. Thirty two African children were given steroid therapy. Thirty (93.7%) did not respond. Five children deteriorated or died during steroid therapy. Very few patients (4) were given cyclophosphamide and none responded. viii Generally intravenous albumen, diuretics and a high protein diet were not very effective in those patients with severe, clinical disease but were of benefit in milder disease. Indian children taken as a whole, responded well to steroid therapy. Seventy-eight percent of the whole group responded to steroids and 21.4% developed cushingoid features. Of the 19 Indian children (all MCNS) treated with cyclophosphamide 63.2% responded of whom about a quarter got toxic side effects (alopecia, darkened nails and leucopenia). Chlorambucil therapy in 4 children (all MCNS) was successful in all. 3.6 Complications Serious infections (septicaemia, peritonitis, urinary tract infection, meningitis, arthritis, osteitis, measles, chicken pox) occurred in 8.7% of the African patients. Eighteen percent had less severe infections. Just over a quarter of the Indian children suffered severe infections. The majority of these patients were MCNS and about 50% were on steroids or cyclophosphamide at the ix time of their infection. Renal biopsy complications were minor, these being abdominal pain and tenderness or transient haematuria. A few patients developed renal haematomas which were detected or monitored by ultrasonography. The single serious complication was the development of a renal abscess at the biopsy site requiring partial nephrectomy. 3.7 Mortality The overall mortality was 5.8%. Seven of the 10 deaths were African children in the Proliferative Group and 3 of the 10 deaths were Indian children. 4. Secondary Nephrotic Syndrome The secondary nephrotics formed an interesting group of patients. Of the 22 patients classified as secondary nephrotics 11 (50%) were related to streptococcal infection either as rapidly progressive glomerulonephritis or transient NS following APSGN. HBsAg was detected in the blood of 8.6% of the African patients. However the HB sAg carrier rate in this age group is 7.4%.The incidence in these patients probably reflects the high incidence in this population group. Collagen vascular disease occurred in 2 patients, both Indian. 5. Conclusions and Recommendations The results of this study demonstrates the strikingly different incidences of the various histological categories in the two race groups studied with a less favourable prognosis and fewer remission rates being achieved in African children. Indian children had more serious infections more often than African children. Steroid and immunosuppressive agents were of no value and probably hazardous in the African child. Some patients deteriorated on these drugs. Indian children who had an excellent response to these drugs were however at significant risk of developing serious infections. Why African children in Durban develop obvious glomerular lesions has not been established. Known or possible aetiological agents such as malaria, schistosomiasis, streptococcal infections and collagen diseases have been excluded. The answer to the above question may in fact lie in genetic predisposition,host factors and environmental influences, either singly or in combination, predisposing to the development of obvious x i glomerular lesions. These require more intensive investigation and judging from the yield of similar studies in other areas of the world expectations have to be guarded.
Human immunodeficiency virus-1 infection and the acquired immunodeficiency syndrome in African children : natural history from birth to early childhood.
(1999) Bobat, Raziya Ahmed.; Coovadia, Hoosen Mahomed.
Background: in 1987, the first child with HIV-1 infection was identified in the paediatric wards at King Edward VIII Hospital in Durban. This made paediatricians aware that the epidemic had spread to the children of KwaZulu/Natal. Although information on transmission and natural history was becoming available from developed countries, little was known about the disease in developing countries. It was important to determine transmission rates and disease patterns in the local population, in order to appropriately counsel women, and for management of infected infants. In addition, with resources for laboratory diagnoses being limited in developing countries, much emphasis had to be placed on clinical findings for identification of infected children. In 1989, a retrospective analysis was made of the HIV-infected children seen over a 2-year period, between 1987 and 1989. Nine such children were identified and their clinical and biochemical features were described. It was concluded that HIV infected children presented with an identifiable pattern of signs, fairly similar to that described for children in industrialised countries. With these findings, a prospective study was undertaken, to determine the vertical transmission rate, the factors affecting this rate, and natural history of vertically transmitted I-IIV-1 infection. ix KwaZulu/Natal, being at the epicentre of the epidemic in South Africa, was a natural site for the study. Patients and Methods: a trained research worker was placed in the antenatal clinic at King Edward VIII Hospital for the specific purpose of educating, counselling, and testing of all women attending the clinic. Women attending the clinic for the first time in the index pregnancy were offered HIV testing if informed consent was obtained. Blood for HIV serology was drawn at the same time as sampling for the obligatory syphilis serology. The acceptance rate for sampling was > 95%. The majority of the women attending the clinic were black, and first attendance was generally late, into the third trimester. The same research worker was responsible for post-test counselling which was offered to all the women, not only those who tested positive. This research worker was also responsible for obtaining maternal consent for entering the newborn infant into the study. All newborn infants were seen within 48 hours of birth. At this time they were examined, growth parameters were recorded, and initial blood samples taken. These infants were then followed-up at 1 month, 2 months, 3 months, then at 3-month intervals up to 18 months, then at 6-month intervals. At each visit, a thorough clinical examination was performed, growth measurements taken, and development assessed. Record was made of any interim illness and visits to health centres, and of hospital admissions. Method of feeding was note& and details on immunisation obtained from the child's immunisation card. The children received all the x routine childhood immunisations according to the national regimen, based on WHO recommendations. Mothers were asked to bring the child to the follow up clinic for any problem, so that episodes of illness would not be missed. The women were reimbursed for transport costs to encourage follow up visits. Calculation of transmission rate and classification of infection status were made according to the recommendations of the Ghent workshop. Children were regarded as infected if they were antibody positive at 18 months or had an HIV related death. They were classified as uninfectd if the antibody test was negative at 9 months of age. Those infants who were lost to follow up before the age of nine months whilst still antibody positive and those whose cause of death could not be determined, were classified as indeterminate. The diagnosis of AIDS was based on the WHO criteria. Blood samples were taken at birth, at age one and three months, then at three month intervals to 18 months; thereafter at six month intervals. Sera were tested for HIV1 antibodies by a commercial enzyme-linked immunosorbent assay,ELISA. Samples that tested positive were confirmed by two tests, a Roche Elisa and by an immunoflourescent assay (IFA). A sample was regarded as being positive if both the second ELISA as well as the IFA or the Western Blot tested positive. xi Results: between October 1990 and March 1993, 234 infants and their 229 mothers were entered into the study. Those who did not attend a single follow up after birth were excluded from the study. The final cohort comprised 181 infants, of whom 48 were classified as infected ( including 17 deaths); 93 not infected, and 40 as indeterminate ( including 8 deaths). Maternal Data: about 60% of the mothers were under 30 years of age and were multiparous; 18% tested positive for syphilis serology; 22.9% were anaemic during pregnancy, and 37% were delivered by caesarean section. Most women lived in urban areas, and 16% chose to bottle-feed exclusively. Vertical Transmission Rate and Factors affecting this Rate: the median vertical transmission rate was 34%, (95% confidence intervals, CI 26%-42%). This figure is similar to that found in most parts of Africa, but much higher than those for Europe and USA. The maternal factors found to be associated with an increased risk of transmission were vaginal deliveries and a low haemoglobin level during pregnancy. Breastfeeding, Transmission, and Outcome: breastfeeding was found to have an increased risk of transmission, by 15 % (CI 1.8-31.8). On assessing growth and morbidity, it was noted that breastfed infants were not protected against such common childhood infections as pneumonia and diarrhoea, and that failure to thrive occurred with equal frequency in both those breastfed as well as those receiving artificial feeds. Newborn Data: when comparing newborn data between those infants who were subsequently found to be infected with those who were uninfected, it was found that there were no major differences between these groups with regard to growth parameters and neonatal complications. However, those infants with rapidly progressive disease (those who died within 24 months), were noted to have lower mean birth weights and lengths, a higher frequency of low birth weights, and tended to have more neonatal problems. Clinical Manifestations: the first differences between the infected and the uninfected infants generally manifested from about 3 months of age. HIV infected children were identifiable by higher frequencies of thrush, lymphadenopathy, skin rash, and hepatosplenomegaly in the early stages, and later on with a higher tendency to neurological and developmental abnormalities, as well as of diarrhoea. Pneumonia was found with equal frequencies in both the infected and uninfected children. The HIV infected child could be distinguished fairly early in life by the combination of the manifestations described above. Progression to AIDS: AIDS was diagnosed in 44% of all the infected children during the study period. Ninety five percent of these children were identified by 12 months of life, showing a rapid progression of the disease Longitudinal Growth: when longitudinal growth parameters were analysed in this cohort, it was found that HIV infected children were stunted from as early as 3 months of age, and remained below the international standards into early childhood. Infected children were also found to be malnourished (i.e. weight for age below international means), from an early age, and this persisted throughout early childhood. Of note, the uninfected childrens' weights, although comparable to international means initially, dropped after the first year of life. However, both groups did not have significant wasting, when compared to international means. Mortality: there were 25 known deaths during the study period. Of these, 17 were classified as HIV-related, and 8 as indeterminate. The mean age at death was 10.1 months, with 83% of all the HIV-related deaths occurring within the first year of life. The commonest diagnoses at the ti me of death were diarrhoea, pneumonia, and failure to thrive; also, thrush was common, as were neurological abnormalities.
Host allergic response variation in children with measles infection.
(1977) Coovadia, Hoosen Mahomed.; Smythe, P. M.
In many infections some patients recover while others die or are permanently disabled. These extremes in clinical outcome may be determined as much by the capacity of the host to eliminate the infecting agent as by the antigenic load on the individual. Children with measles who do not recover may succumb to acute complications (mainly respiratory) or chronic disease (respiratory and neurological) may develop. Analysis of immunological function antedating any of these final events would assist in understanding their pathogenesis and possibly aid in management. In order to achieve t h i s , immunological responsiveness was at f i r st studied in 24 children with acute measles and compared with that in 20 children with established chronic post measles chest disease investigated 6 - 1 6 weeks after appearance of the rash. The immunosuppressive effects of acute measles were extensive. Total white cells were reduced and this reduction was accounted for entirely by lymphopenia which was equally expressed among the major lymphocyte sub-populations studied; the function of T cells, assessed by radioisotope incorporation into phytohaemaggiutinin-transformed lymphocytes and by delayed skin hypersensitivity to dinitrochlorobenzene, was depressed. Serum IgA was reduced in acute measles patients. In contrast there was a relative sparing of the measured indices of immunity in patients with chronic post measles chest disease, with the major defect being an impaired delayed hypersensitivity reaction to dinitrochlorobenzene. There were minor alterations in complement components in both groups of patients with the evidence suggesting minimal utilisation of the alternative pathway in acute measles and classical pathway in chronic patients. High levels of heterophile antibodies to sheep red blood cells were detected in patients with chronic chest disease. (11) The results suggested that the conditions for chronicity of pulmonary disease in measles were unlikely to be determined by persistent abnormalities in the immunopathological factors enumerated, most of which were normal in chronic patients. It was not possible to interpret the findings of defective delayed hypersensitivity and complement components in patients with chronic chest disease as being either the cause or the effect of chronicity. The latter findings would have to be compared with results in children who had recovered from measles studied six weeks after onset of rash. An attempt was made to resolve this problem. Twenty-two children with measles were studied in the acute stage of the rash and six weeks later and results compared with matched controls. The above findings in acute measles were confirmed: the total lymphocyte count and major lymphocyte sub-populations were significantly below control values. At six weeks the B cell and Null cell counts were s t i ll significantly diminished. The function of T cells assessed by radioisotope uptake by phytohaemagglutinin-stimulated lymphocytes and by delayed skin hypersensitivity reaction to dinitrochlorobenzene was impaired during the acute stage and this persisted for six weeks. No important abnormalities were detected in serum immunoglobulins and complement components. Partial reversal of immunological suppression caused by measles was therefore demonstrated at 6 weeks after the appearance of rash. Demonstration of a persistently defective delayed hypersensitivity in those who recovered made i t unlikely that this anergy was important in the development of chronicity. Complement abnormalities were similarly unrelated to progression to chronic lung damage. ( Children who recovered, when studied at six weeks, appeared to be worse o f f immunologically than those with established chronic chest disease following measles. Children with chronic chest disease were studied 6 - 1 6 weeks after onset of rash, by which time the partial reversal of immune deficiency, noted at 6 weeks, would be complete. Among the group of children studied during the acute rash of measles there were five who subsequently died and one who progressed to chronic chest disease. Results in these six children were compared with those in six age-matched children who recovered from measles within a week. In the children who subsequently died or developed chronic pneumonia, immunosuppression was more pronounced during the acute rash ( i . e ., 3 - 2 0 days before death) than in the children who recovered. The absolute total lymphocyte count (T and B cells) was s i g n i f i c a n t ly lower in those who died or developed chronicity. Mean serum C, was also lower in this group. There were no significant differences between the two groups for total white c e l l s , neutrophils, Null c e l l s, cells with both T and B cell markers, other complement factors, serum immunoglobulins and phytohaemagglutinin stimulation of lymphocytes. The total lymphocyte count in a further nineteen patients with measles who had died, studied retrospectively, was s i g n i f i c a n t l y lower than that in twenty-seven patients with measles who recovered. Children whose outcome was poor generally had absolute lymphocyte 3 counts below 2000 cells/mm whereas those who recovered had values above this level. (iv) Therefore children who w i l l die or develop chronic chest disease can be often distinguished, within two days of the appearance of the rash, from those who w i l l recover. In order to test the v a l i d i t y of this conclusion based on results obtained from a small sample the study was extended so as to increase the number of patients with measles who had severe lymphopenia (< 2000 cells/mm3). Seventy seven per cent of 30 children who had severe lymphopenia within 2 days of appearance of rash f a i l e d to recover: 30% died from pulmonary complications within a few days to two months of the onset of the exanthem while 47% developed chronic lung damage. This was s i g n i f i c a n t ly worse than the outcome in 30 children with lymphocyte counts above 2000 3 cells/mm , of whom 67% recovered, 33% developed chronic chest disease and none died. Persistence of severe lymphopenia (which was due to reduction 3 in both T and B cells) in those with i n i t i a l counts below 2000 cells/mm , for at least fifteen days after onset of rash, remained a good predictive index of morbidity and mortality. Reversal of immunoparesis in those with i n i t i a l severe lymphopenia was slower and less complete 42 days from the appearance of the rash in children who subsequently died or progressed to chronicity than in those who recovered. All patients who died f a i l ed to produce an adequate or sustained antibody response to measles. The results of these studies suggest that long term pulmonary and possibly neurological sequelae of measles are probably due to a transient widespread immunoparesis during early measles with persistent defects in specific immunity to measles and probably other viruses, whereas recovery is due to less severe effects of shorter duration. (v) In order to answer the question why some children do badly and others well after measles, studies on the HLA frequencies and measles antigen load have been undertaken in children with severe lymphopenia. Results of viral load are inconclusive and those of HLA suggest a trend towards histocompatibility linked genetic susceptibility to the development of severe lymphopenia in measles associated with HLA AW32. The therapeutic implication of these studies is that children with measles who are at risk for death and chronic disease can be identified early in the disease and intervention at this stage may reverse the severe immunosuppression which leads to rapid demise or modify the immunopathological changes progressing relentlessly in some cases to permanent lung and brain damage and occasionally to death.
Aids for the early diagnosis of tuberculous meningitis (TBM)
(1985) Ramkissoon, Arthi.; Coovadia, Hoosen Mahomed.
Mortality and morbidity rates associated with tuberculous meningitis (TBM) are substantial. The average duration of the untreated disease from onset to death is about 17 days. The prognosis of TBM is known to correlate with the stage of the disease at the time of diagnosis and commencement of chemotherapy. Early diagnosis improves the chances of recovery without neurological sequelae. Early diagnosis is a problem because the presenting symptoms are non-specific and the onset of the disease is typically insidious. To date no single test is available that is totally reliable and specific for TBM. I have attempted to develop a reliable and easily applicable test for the diagnosis of TBM. In fulfilling this objective, the work undertaken may be divided into three major sections:- 1. Detection of soluble Mycobacterium tuberculosis antigens in the cerebrospinal fluid (CSF) of patients with TBM and in control groups by using Mycobacterium bovis BCG antigens. The technique used was that of inhibition enzyme-linked immunosorbent assay (ELISA). The principle of this technique is illustrated in Fig. 5. 2. Detection of soluble M. tuberculosis antigens in the CSF of tuberculous and control groups of patients by using antibodies raised against M.bovis BCG. The technique used was that of the double antibody sandwich ELISA. An outline of this ELISA is given in Fig. 6. 3. Correlation of chloride levels in the blood and CSF of patients with tuberculous and other forms of meningitis. It has been established that the SERUM/CSF ratio of bromide tends towards unity in patients with TBM because the permeability of the blood-brain barrier is impaired. Since both bromide and chloride are chemically similar (both being halides), it was thought that a similar pattern may exist for BLOOD/CSF chloride ratios; and this was investigated. The method used for the INHIBITION ELISA had to be standardized before the samples could be tested. This involved investigating the acceptability of various microtitre plates; determination of the optimal working dilutions for the coating solution and conjugate; and determination of optimal conditions for the various incubation periods, both in terms of time and temperature. A total of 70 specimens was tested. These consisted of 25 normal CSF controls; 25 pleural and ascitic fluid samples; 10 TBM samples, and 10 bacterial meningitis CSF samples. It was found that a distinction existed between the absorbance values obtained from positive TBM CSF samples (Mean 0,658 + 0,043) and that from normal CSF samples (Mean 1,089 + 0,224). The mean absorbance of the culture-positive bacterial CSF's also differed significantly from the other 2 groups (Tables VII; IX). Some overlap occurred amongst the absorbance values of bacterial culture positive CSF's (Range 0,975-0,879) and normal CSF's (Range 1,486-0,934). The mean absorbance value for bacterial positive CSF samples (0,920 _+ 0,029) differed significantly (p <0,01) from those of normal CSF (1,089 + 0,224) and TBM CSF's (0,658 + 0,043). The difference between the mean values obtained with tuberculous and non-tuberculous groups of pleural and ascitic fluid was also significant (p < 0,01). The method used for the DOUBLE ANTIBODY SANDWICH ELISA was that of Sada et al. (1983). Before the samples could be tested, the method had to be standardized and similar investigations to those for the INHIBITION ELISA were performed. In addition, antibodies raised against M.bovis BCG were conjugated to alkaline phosphatase since no commercial preparation was available. Unfortunately no distinction was recorded between negative and positive test specimens, even on repetition of the entire procedure. Measurement of chloride was done by a fully automated procedure using the BECKMAN ASTRA-8. A total of 149 samples were tested. Of these 10 were tuberculous, 34 were viral, and the remainder were bacterial meningitis. No pattern was established that could differentiate TBM from viral or bacterial meningitis. The results obtained are tabulated in Table III and illustrated in Figures 9, 10, and 11. In summarizing, the use of the INHIBITION ELISA technique for the accurate diagnosis of TBM seems promising. However, its validity in the clinical situation will have to be assessed further and with greater numbers of specimens before it can be adopted as a diagnostic procedure for TBM. OBJECTIVE. To determine 1. The ability and reliability of the INHIBITION ELISA1 technique to detect mycobacterial antigens in pleural, ascitic, and cerebrospinal fluids. 2. The accuracy and reproducibility of the double antibody sandwich ELISA in the detection of mycobacterial antigens in CSF of patients with tuberculous meningitis (TBM). 3. Whether a correlation exists between blood and CSF chloride levels in patients with tuberculous and other forms of meningitis.
Assisted respiration in the treatment of neonatal tetanus.
(1967) Thambiran, A. K.
The effect of corticosteroid therapy on growth in Black South African children with nephrotic syndrome.
(1986) Manikkam, Noel Edwin George.
The most useful drugs in the management of nephrotic syndrome are the corticosteroids. These drugs are as well known for their adverse effects as they are for their therapeutic advantages. The two most common paediatric side effects are suppression of linear growth and posterior subcapsular cataracts. Both of these untoward effects are insiduous and therefore less easily perceived. Although many workers have studied the growth inhibiting effects of the corticosteroids in the various diseases e.g. asthma, very little work was done to investigate these effects in patients with nephrotic syndrome. Furthermore, the Renal Clinic, King Edward VIII Hospital, Durban continues to use a daily regime of prednisone instead of the alternate day regime which is widely recommended to minimise growth retardation. This study was therefore undertaken to investigate the growth inhibiting effects of repeated courses of daily, high-dose prednisone in African and Indian children with nephrotic syndrome. All children with nephrotic syndrome with relevant data in their records and with no other chronic illness were selected from the Renal Clinic. Of the 125 selected, 87 children had been treated with prednisone for an average of 35,9 weeks and 38 had been treated symptomatically. The heights of those that received prednisone were measured at an averace of 77 weeks after completion of therapy. The mean height standard deviation score (SDS) of the treatment and control groups of Indian children were -1,06 and -0,92 respectively, both being between the 10th and 25th percentile, whilst the mean height SDS of the treatment and control groups of African children were -1,82 (just below the 5th percentile) and -1,77 (between the 5th and 10th percentile) respectively. From the results, it is evident that repeated courses of daily prednisone therapy, even when it exceeds 36 weeks, does not inhibit growth in both African and Indian children. Although there was no significant difference between the races and sexes with respect to growth and corticosteroid therapy, this study does confirm earlier reports that most of the African children with nephrotic syndrome had obvious glomerular lesions whilst most of the Indians had minimal change nephrotic syndrome.
The interaction between human leucocyte antigen-G and natural killer cells at the placental interface in HIV-1 infected pregnant women and the significance, if any, to in utero transmission.
(2007) Moodley, Shamala.; Carpenter, S.; Bobat, Raziya Ahmed.
This study was undertaken to investigate the relationship between Natural Killer cells and HLA-G at the placental barrier in HIV-I infected pregnant women and to establish the significance, if any, to in utero infection. Fifty-five HIV -I infected pregnant women were recruited into the study after consent was obtained. Blood samples were collected from both mothers and babies for viral loads and CD4+ cell counts. Placental samples were obtained from pregnancies at delivery and examined by immunoperoxidase immunohistochemistry methods using monoclonal antibodies to p24 antigens and Natural Killer (CD56+) cells. HLA-G expression was quantified using real-time polymerase chain reaction. Analysis of viral loads and CD4+ cell counts were undertaken in categories. No significant association was observed between the viral load of mothers and their CD4+ cell counts. Eighteen percent of the women in this study population had 5 log viral loads with a transmission rate of 0.27(95% Cl, 0.15 - O. 39). Maternal viraemia was significantly associated with transmission of infection to babies (p = 0.047). The odds ratio indicated that for every 1 log increase in maternal viral load the babies were 3.1 times more likely to acquire the infection (Exp (B) = 3.137 (95%CI, 1.015-9.696). Furthermore, the study found that a higher number of female babies were infected than males. Although not statistically significant the odds ratio indicated that female babies were 3.1 times more likely to become infected than males (Exp (B) = 3.110 (95%CI, 0.819-11.808). We report here the results of immunohistochemistry for p24 antigens and NK (CD56+) cells and compare them to the immunological responses of both mothers and babies at birth. HIV-1 antigens were detected in 94.5% of all placentas by immunohistochemistry. Infiltration of CD56+ was found in 98% of placental tissue. The analysis revealed that the presence of p24 antigens in placental tissue was not influenced by maternal viral load or CD4+ cell counts. Lower median NK cell values were observed in placentas of mothers with infected babies as compared with the uninfected cluster. Although not statistically significant, the risk of vertical transmission was increased 3.4 times more in placentas which had lower NK cell values. According to the odds ratio, babies CD4+ counts were affected by every 1 log increase in mother's viral load. Overall, maternal viral load emerged as a strong predictor for risk of infection from infected mothers to their infants. Our analysis indicated that female babies were 3.7 times more likely to acquire the infection than males. Using data obtained from real-time PCR we investigated the relationship between maternal viral load and the quantity of HLA-G expression (p = 0.045; 95%CI 1.029- 11.499). Logistic regression models revealed that mother's viral load was the strongest risk factor for vertical transmission. No statistically significant correlation was noted with HLA-G and viral transmission. However, the odds ratio indicated that the risk of infection increased by 1.3 with every 1 fold increase in HLA-G expression. An analysis of mother-to-child transmission rates by gender revealed that the odds ratio for transmission was 3.4 times more in female babies than in males. We then investigated the relationship between maternal viraemia and HLA-G expression. A positive correlation between maternal viral load and placental HLA-G was observed (p = 0.038). When gender susceptibility to HLA-G expression was explored a statistically significant association was observed in placental tissue of mothers with infected and uninfected male babies and HLA-G expression (p = 0.013). To conclude, the analysis found that HLA-G was up regulated 3.95 times more in placental tissue of mothers with infected babies than in mothers with uninfected babies.
HIV-1 specific T-Cell responses in chronic HIV infected children during continuous treatment and structured treatment interruptions (STI).
(2010) Reddy, Shabashini.; Ndung'u, Peter Thumbi.
BACKGROUND Sub-Saharan Africa has the highest number of HIV-infected individuals and limited treatment programs. The use of Highly Active Antiretroviral Therapy (HAART) has resulted in a considerable decrease in morbidity and mortality among HIV-infected individuals. Long-term use of HAART has several limitations relating to cost, drug toxicity and adherence. Structured Treatment Interruption (STI) has been proposed as a therapeutic approach which limits the exposure to continuous HAART, but retains the benefits thereof. The role of HIV-specific Tcell responses in the control of viraemia has not been well studied in children and it is not clear when these responses become detectable or whether they are associated with improved viral control. Furthermore, antiretroviral drug resistance is well documented in adults infected with HIV-1 clade B virus but comparable information is lacking for chronic paediatric clade C virus infection. This pilot study focused on a chronic HIV-infected paediatric cohort from Durban, South Africa, to assess the immunologic and virologic responses in perinatal HIVinfected children undergoing STI. METHODS Thirty chronic HIV-infected treatment naïve children were enrolled and randomised into either the treatment interruption or continuous treatment group. Longitudinal measurements of viral loads and CD4 percentages were done at scheduled intervals. Peripheral blood mononuclear cells (PBMCs) were screened for cytotoxic T-lymphocyte (CTL) gamma interferon (IFN-?) enzyme-linked immunospot (ELISpot) assay responses using 410 peptides which spanned the entire HIV-1 clade C proteome. Intracellular cytokine staining (ICS) was done to distinguish between IFN-? Gag-specific T-helper and cytotoxic T cell responses. Pre-HAART drug resistance mutations testing and HLA typing were done for all children. RESULTS There was a significant increase in the median CD4 percentage after HAART was introduced. Six children randomized to the STI arm did not undergo treatment interruption because their viral loads remained detectable at the time of scheduled interruption. Most HIV proteins were targeted in this paediatric cohort. Gag was the most frequently targeted HIV-1 protein (93.1%). In both treatment groups, there were broadening of T-cell responses, however, the magnitude of T-cell responses decreased over time on HAART. Drug-resistant mutations were detectable in 4/29 children before initiation of HAART. CONCLUSION In this pilot study, the HIV-1-specific CD8+ and CD4+ T-cell responses were detected before and during HAART. Although the treatment interruption period was short, there were no adverse outcomes in either the continuous or treatment interruption groups in terms of death or other clinical outcomes. This study suggests that it is important to continue to explore alternative treatment strategies in order to reduce cost and toxicity as well as to enhance adherence.
Epidemiological and clinical studies of vitamin A in Black South African pre-school children.
(1993) Coutsoudis, Anna.
The ocular complications of vitamin A deficiency have been known for many years, however, recent studies have suggested that marginal vitamin A status enlarges the risk of common childhood infections and increases mortality. It is therefore important to assess the vitamin A status, and some of its consequences, in children who are most likely to be at risk for vitamin A deficiency as this has important implications for promoting the health of children and for formulating appropriate primary health care policies. In South Africa very little data is available on vitamin A nutrition of communities; therefore one of the objectives of this research programme was to document the vitamin A status of African children who, because of historical inequities, are most likely to be at risk for deficiency. Sound, epidemiologically based surveys of vitamin A intake and body levels were conducted in a typical established township (using dietary intake as the measuring tool) and in a typical peri-urban informal settlement (using serum retinol and conjunctival impression cytology as the measuring tools). These studies revealed that the majority (97%) of children living in the established township surveyed had an adequate intake of vitamin A, whereas 44% of the children in the informal settlement had low serum retinol levels (20 ug/dL), and 18% had insufficient vitamin A, as assessed by 2 abnormal disc specimens, using the conjunctival impression cytology test. In order to investigate the interrelationsnips between vitamin A, other micronutrients and some risk factors, an analysis was undertaken of anthropometry, parasite infestation and blood concentrations of vitamin E, calcium, magnesium, phosphorous, albumin, haemaglobin, serum iron and ferritin and percent transferrin saturation. Significant positive correlations were found between serum retinol and all the biochemical indicators of iron metabolism studied except for serum ferritin. Ninety one percent of the children sampled were infested with parasites. These results highlight the fact that in this population close interconnections exist among nutrients and suggest that attempts at correcting vitamin A deficiency in such communities should be based on comprehensive intervention programmes rather than on single nutrient replacement. The impact of infections on blood levels of vitamin A was investigated in African children with severe measles. In addition, substances related to vitamin A metabolism such as other micronutrients (zinc, vitamin E) and proteins (retinol binding protein, prealbumin, albumin) were measured in serum. In addition the changes induced in these substances by vitamin A supplementation (offered in a randomised, double blind, placebo controlled trial) were studied. Serum retinol as well as the other nutrients measured were significantly reduced early in the exanthem in measles patients as compared to healthy controls. Vitamin A and prealbumin levels on day 8 (of the intervention trial) were significantly increased in the supplemented group compared to the placebo group. vitamin A levels in serum correlated with those of retinol binding protein (RBP), prealbumin and zinc. These findings strengthen the hypothesis that hyporetinaemia during measles is the consequence of impaired mobilisation of retinol stores from the liver. The effect of reversing the temporary lowering of serum retinol concentrations during acute measles infections by supplementation with vitamin A was investigated in a hospital based, randomized, double-blind, placebo controlled trial. The two groups were comparable in known covariates of measles severity : weight/age centiles; overcrowding; rash; total 90% of the patients had blood lymphocytes; serum levels pre-albumin, RBP, vitamins A and E. of zinc, albumin, hyporetinaemia. Integrated Morbidity Scores ( IMS) derived from diarrhoea, herpes and respiratory tract infection (radiologically confirmed) were assigned on day 8, at 6 weeks and 6 months - these were reduced by 82%, 61% and 85% respectively in the supplemented group. This was mainly due to reduced respiratory tract infection. There was one death in the placebo group. At 6 weeks there was significant weight gain in the supplemented group. Despite the selected sample, attention to multiple covariates enhances the validity of the data obtained and supports the current WHO recommendations for vitamin A supplementation during measles. There are several mechanisms by which vitamin A is thought to have its effect of reducing morbidity, one of which is by improving immune responsiveness. This particular mechanism has not been adequately studied in children; most of the studies having been conducted in animals. The effect of vitamin A supplementation on selected factors of immunity in African children with complicated measles was therefore investigated during the randomized double-blind, placebo controlled, intervention trial described above. Placebo and treated groups had similar baseline characteristics. In the treated group there was a significant increase in total number of lymphocytes (day 42, P = .05) and measles IgG antibody concentrations (day 8, p = .02), both of which have consistently been shown to correlate more closely with outcome in measles than other immunological, clinical and radiological factors. Interleukin-2 (IL-2) and plasma complement (C3 ) values were unaffected by vitamin A supplement.at.Lon , These findings reinforce results from animal studies which show that the pathways of vitamin A activity in decreasing morbidity and mortality are partly founded on selective immunopotentiation. In conclusion epidemiological and biochemical methods which were used to assess the vitamin A status of African children in South Africa revealed that overt vitamin A deficiency is not a Public health issue to the extent it is in the poor rice eating nations of the world. Marginal vitamin A deficiency is however prevalent in informal settlements. Interventions to reverse this marginal vitamin A deficiency should be incorporated in comprehensive programmes to ensure food security. Infections such as measles which increase utilisation and inhibit mobilisation from body stores are damaging to vitamin A homeostasis in the individual. The morbidity associated with measles can be reversed by high dose vitamin A supplementation during the acute phase of the infection. Improving immune responsiveness is one of the likely paths of vitamin A activity in decreasing morbidity from measles.
Investigation into optimal amikacin dosing in children.
(1996) Forsyth, Nan Barbara.
Aminoglycoside antibacterial agents, such as amikacin, continue to play an important role in the treatment of Gram-negative infections. However, although extremely effective, they are not without potential adverse events, the most important of which being nephro- and ototoxicity. Research into factors thought to influence both the efficacy and toxicity, has challenged the rationale upon which these agents have classically been dosed. Various studies in adult patients have found that a new approach to dosing (use of single daily administration) has equal or greater efficacy or safety compared to the standard multiple daily dosing of these agents. Similar studies comparing regimens in children are few, and as yet no comparative investigation has been performed using amikacin in children (as a separate and distinct group). Additionally, in evaluating the impact of altering dose regimens, it is imperative that the documented age-related aminoglycoside pharmacokinetic alterations, be taken into account. Amikacin pharmacokinetic parameters (determined using traditional methods) have been previously published for various (usually small) groups of children. However, population parameters are not currently available for South African children . This study therefore aimed to investigate optimal amikacin dosing in children by studying: a) the comparative efficacy and toxicity of two dosing regimens, and b) the population pharmacokinetic parameters derived using one of the alternative approaches capable of utilising routine, sparse serum drug concentration time data. This investigation was conducted in the paediatric surgical and burns wards of King Edward VIII Hospital , Durban. Study patients (0.6-12 years) received amikacin either once daily (15mg/kg) or twice daily (7.5 mg/kg) by slow intravenous bolus. Concomitant medication was given as prescribed. Regimen efficacy (favourable, unfavourable or indeterminate outcome) was assessed by patient temperatures, clinical improvement and white cell counts. Clinical nephrotoxicity was evaluated by changes in serum creatinine, and renal tubular damage (investigated in a small subgroup of patients) was indicated by detection of urinary low molecular weight proteins. Ototoxicity (cochleotoxicity) was assessed by pure tone audiometry. Pertinent demographic and treatment details (amikacin concentration time data) were recorded for the population pharmacokinetic analysis. The Nonlinear Mixed Effects Model (NONMEM) programme was used to derive appropriate models describing clearance (CL) and volume of distribution (V), as well as mean values of these pharmacokinetic parameters for this population. Fifty four patients were entered into the regimen assessment. Patients in the single daily regimen (n=27) had significantly greater (p<0.05) mean (SO) peak (±0.5 hour post-dose) serum amikacin levels (37.7 (6.9) mg/L) as well as cumulative dose (91.5 (26.5) mg/kg) and duration of therapy (5.7 (1 .5) days) when compared with those of the twice daily group (19.5 (3.7) mg/L, 70.1 (26.1) mg/kg and 4.6 (1 .6) days respectively). No statistically significant differences were found between the groups in terms of outcome (18/24 and 22/25 patients in the once and twice daily dosing groups had favourable outcomes; there were no unfavourable outcomes). Pure tone audiometry (evaluated post-therapy , in 20 patients from each dosing regimen) revealed no statistically significant differences between the number of patients in the two groups with possible drug-related ototoxicity. None of the patients assessed (including an additional 14 patients with burn injury) developed clinical nephrotoxicity. Urinalysis was performed in 17 amikacin treated patients (9 and 8 from the once and twice daily dosing regimens respectively) and 9 control subjects. Low molecular weight proteinuria was absent in all of the latter patients except one, in whom pre-existing renal disease was suspected. Tubular dysfunction ascribed to possible drug effect was detected in similar numbers of patients in the two treatment groups (3 and 2 patients in the once and twice daily dosing groups respectively). In the pharmacokinetic assessment (156 serum levels from 82 patients) using a one compartment model, the final models which best described the data were as follows : CL (Uhr) = 0.271 x age(yrs) + 2.46 x body surface areatrrr'), V (L) = 7.34 x body surface areatrn") Other fixed effects tested, which did not render the data more probable, included serum creatinine measurements at the start of treatment, gender, presence of burn injury and drug regimen. Interpatient variation was 15% and 18% for CL and V respectively, with intrapatient variation or residual error of 10%. The weight adjusted population parameter estimates (95% Confidence Interval) for this group were CL =0.180 (0.175 ,0.185) Uhr/kg and V =0.293 (0.286, 0.300) Ukg, which are within the range of values published previously for other children of similar ages. The findings of this investigation , consistent with those of other similar studies, indicate that daily amikacin administration (in combination with a B-lactam), to children with normal renal function, has similar efficacy to, and no greater toxicity than multiple daily dosing. However, the role, if any, of the significantly greater cumulative dose and duration of therapy in the daily dosing group is unknown. As uncertainty remains regarding the precise duration of certain post-exposure events (and hence, the ideal duration of the interdose interval), and with the rapid drug clearance in this group of patients , future in vitro and in vivo investigations may shed even further light on the optimal dosing approach in these patients.
Pertussis vaccination of African infants using acellular and whole-cell vaccines.
(1991) Ramkissoon, Arthi.
Conventional pertussis vaccine prepared from killed whole cell B. pertussis organisms has been in widespread use since the early 1950's. Despite marked reductions in the incidence of pertussis, the use of the vaccine has caused concern because of questions of significant adverse reactions. Whooping cough is not notifiable in South Africa, and there is consequently a paucity of hard data on efficacy; in addition few cases are proven. Incidence, prevalence, severity and transmission of the disease hence remain a matter of conjecture. In order to provide background information and determine baseline data for undertaking further studies, available clinical and epidemiological data on whooping cough (pertussis) in South Africa was collated. It was intended to compare the pattern of disease seen in this country with that known in other parts of the world. Clinical and epidemiological findings from 1525 whooping cough admissions (diagnosed on the basis of clinical criteria) obtained from 6 major infectious disease hospitals around the country for the period 1980-1987/1988 are reported. The data from Durban were available in some detail. Incidence, morbidity and mortality of the disease in South Africa, as elsewhere, was higher in infancy. Infants and young children were predominantly affected, with 31.3% of cases under 12 months of age and 7.2% less than 2 months. Mortality was disproportionately higher in infancy; 53 .2% of deaths were in those younger than 12 months of age. There was a slight female preponderance, both in respect of prevalence and mortality. Patients were admitted with pertussis throughout the year, although there was a peak during the winter months (May to October). The typical whoop accompanied the cough in 55.6% of patients . A raised white cell count was recorded in 66% of patients. The most commonly detected The average duration of hospital stay was between 1 0-13 days. Of those with vaccination records, 26% were unvaccinated, 44% had 1 or 2 doses, and 27% had been fully vaccinated with a whole-cell pertussis vaccine combined with diphtheria and tetanus toxoids. The picture, incomplete though it is, reveals a pattern of pertussis similar to that described in other developing countries. The study reveals huge gaps in our knowledge of this subject in South Africa. More research needs to be done, particularly with respect to improved diagnosis, prevention and treatment; further pertussis should be made a notifiable disease in South Africa. The whole-cell pertussis vaccine currently used in South Africa has not been adequately evaluated for post-vaccination events and immunogenicity. The development of new purified component pertussis vaccines, which appear to be safer than conventional whole-cell preparations and of equal or almost equal efficacy (although optimal vaccine composition remains to be defined), requires that the concept of early vaccination with this vaccine compared with conventional whole-cell vaccine be examined in order to optimise the immune response to these vaccines in infants; more especially since neonates do not appear to benefit from passive immunity. Acellular pertussis vaccine has not been evaluated previously in neonates. In order to address the problem of high morbidity and mortality from pertussis in early infancy; and the incorporation of the vaccine into routine vaccination schedules, a phase 11 trial of acellular and whole-cell vaccines was undertaken in very young infants. The effect of neonatal vaccination with acellular pertussis vaccine on subsequent immunity; and the immunogenicity and shortterm safety and reactogenicity of routine primary vaccination with acellular vaccine compared with conventional whole-cell preparations was investigated. Three hundred and forty-five healthy, full-term African babies were, enrolled in the study at birth; 58% of whom were successfully followed for 9 months. Infants were assigned to 1 of 3 vaccine groups in sequence at birth and received either acellular or whole-cell pertussis vaccine , combined with 0 and T (A-OTP or W-OTP) at 2, 4 and 6 months of age. Groups I and 11 received A-OTP and Group III W-OTP. In addition, at birth, Group I received an additional dose of A-OTP and Group 11, a saline placebo injection. No unvaccinated controls were studied for ethical reasons. Serologic IgG responses to 3 major protective antigens of B. pertussis, filamentous haemagglutinin (FHA), pertussis toxin (PT) and fimbrial agglutinogens 2,3 (AGG2,3), were measured by ELlSA in sera obtained at birth, and before vaccination at 2, 4 and 6 months and at 9 months of age. The incidence and nature of post-vaccination events were recorded for 14 days after each dose. A-OTP induced serum IgG responses to PT and FHA comparable with those reported in other studies, with peak PT titres occurring at 6 months of age after 2 doses in babies vaccinated according to the routine schedule (Group 11). Surprisingly, response to W-OTP was found merely to restore levels of antibody to PT and FHA to those found in cord blood after 3 doses of vaccine, which questions the immunogenicity of the South African product. Four doses of AOTP (Group I) did not produce a better antibody response than the 3-dose schedule. Incidence of all post-vaccination events to both acellular and whole-cell vaccines was low (85.96/1000 doses, 136.36/1000 doses and 76.30/1000 doses in Groups I, 11 and III respectively). Minor symptoms were more common in recipients of A-OTP, although no significant differences in rates were demonstrated. Neurologic post-vaccination events (without sequelae) occurred more frequently hi recipients of W-OTP. No infant vaccinated with A-OTP from 2 months of age (Group 11) experienced a neurologic symptom. The risks of major post-vaccination events cannot however be fully quantified In a study of this small size and diseases. A-DTP vaccination commencing at birth produced final antibody titres to PT and FHA which were superior to those of South African whole-cell vaccine but were considerably lower than when the vaccine was incorporated into routine schedules commencing at 2 months of age. The study findings suggest that acellular pertussis vaccines, whether given from birth or from the age of 2 months, appeared safer and produced a better serologic response than the South African whole-cell product which may have impaired immunogenicity. During the course of the above study, 11 full-term infants with pertussis infection (10 subclinical) were retrospectively diagnosed on the basis of ser~logic evidence. Of the infants with subclinical disease, all 10 had a ~ 4-fold rise in at least 2 different pertussis IgG antibodies and nine had ~ 4-fold rise in all 3 IgG antibodies measured. Seven infants had raised IgA antibodies to PT and FHA and 4 infants had raised IgA antibodies to AGG2,3. Subclinical infection provoked antibody production to multiple antigens to differing degrees. The role of various factors which may have contributed to asymptomatic infection were analysed, viz - household contacts; type of antibody response (clinical vs. subclinical; vaccinated vs. subclinical); maternally acquired antibody levels; vaccination status (number of vaccine doses received); age and gender; and nutritional status. SpeCial features of the study which require emphasis are: pertussis remained subclinical in unvaccinated babies (most of the subjects were unvaccinated). Subclinical infection followed incomplete primary vaccination with either acellular or whole-cell vaccines commencing at 2 months of age. Subjects were 8 months of age or younger and only 1 had completed primary vaccination. Other infections of infancy were commonly detected; 4 infants had upper Likelihood of subclinical infection was related to significantly lower levels of maternallyacquired pertussis IgG-AGG2,3 antibodies but not associated with infants' nutritional status. Subclinical pertussis is described in very young African babies at an age when the disease is most severe, and therefore has implications for infant morbidity and mortality; it is also relevant to disease surveillance and vaccine-efficacy studies . Some perinatal factors influencing vaccination were also explored . In this context we looked at: i. The acquisition of maternal antibodies to B. pertussis in paired mother-infant sera from both well -nourished and SGA full-term and pre-term infants, and infants who subsequently developed pertussis infection, and effect of these maternally-acquired antibodies on subsequent antibody responses to acellular pertussis vaccine administered soon after birth, and to acellular and whole-cell vaccines administered from the age of 2 months. if. The acquisition of maternal antibodies to diphtheria and tetanus (OT) in paired motherinfant sera from full-term and pre-term infants, and the effect of these maternallyacquired antibodies on serologic responses to neonatal OT vaccination followed by whole-cell OTP vaccination at 2, 4 and 6 months. Maternal antibodies were measured since little is known about materno-fetal transfer of pertussis antibodies, especially in African countries where inhibition of placental transfer might occur for a variety of reasons. Furthermore, the effect of peri-natal malnutrition and prematurity on transplacental transfer of diphtheria, tetanus and pertussis antibodies has not been conclusively established in these areas. sera with levels in the latter frequently being higher, Indicating active transplacental transfer of antibodies. The significant pertussis antibody levels in maternal sera were unlikely to be due to the currently used South African whole-cell vaccine (given the poor antibody response to PT and FHA shown in this study). It is assumed that the presence of these antibodies are the end result of natural infection and therefore that pertussis is widespread in the African community. Maternal and cord IgG-PT and AGG2,3 titres were significantly lower (p <0.05) and maternal IgG-FHA marginally lower (p ... O.05) in SGA infants compared to cohorts, although placental transfer was equally efficient in both groups~ This study has demonstrated that the high titres of maternally derived circulating B. pertussis antibodies do not have an inhibitory effect on the subsequent serologic response to acellular vaccine administered in early infancy (2 months) or with the first dose given soon after birth. Protective levels of diphtheria and tetanus antibodies were detected in 100% and 76% of cord sera in pre-term infants and in 85% and 67% of cord sera in full-term infants. Although the number of infants studied was too small to make a definite comment, there did not appear to be 'tolerance' due to neonatal diphtheria-tetanus vaccination of pre-term infants, or to high levels of maternally-acquired antibody. During analysis of the data from the phase 11 study of acellular pertussis vacclnes, 25 infants with protein-energy-malnutritlon (PEM) were detected on the basis of anthropometric indices. Seventeen infants were smaIJ-for-gestatlonal-age (SGA). of whom 9 developed PEM by the age of 9 months. Eight other infants developed post-natal PEM before the completion of the primary vaccination course. and extent of immunological impairment, if any, on the serologic responses to acellular vaccine and to conventional whole-cell OTP in SGA infants, and in infants who developed PEM before the completion of the primary vaccination course. The following indices were evaluated in malnourished infants; (i) anthropometric indices of nutritional status, (ii) intercurrent illnesses including pertussis infection, (iii) post-vaccination events, (iv) serologic responses to vaccination. Results were compared with those obtained in well-nourished (WN) age- and vaccine-matched cohorts. Overall, peak titres and seroconversion data of all 3 antibodies were not significantly different in malnourished infants though final anti-AGG2,3 titres (at 9 months of age) in Group III were significantly lower (p = 0.035). Although peak and final PT and FHA antibody titres in many SGA and malnourished infants were lower than in WN infants and peak responses were attained at a later age, malnutrition did not significantly affect the response to A-OTP. Peak and final AGG2,3 antibody titres were similar in SGA, malnourished and WN infants. Overall malnourished infants responded no less well to pertussis vaccination than did other vaccinees. Incidence of minor local and systemic post-vaccination symptoms was not significantly different in PEM and WN groups although induration at injection site and irritability were more frequently reported in the latter. No major neurologic post-vaccination symptoms to either vaccine were reported in SGA infants or infants with PEM at the time of vaccination. No significant differences was noted in the incidence of major symptoms in PEM as compared with WN infants. One male infant (Group I) who was malnourished at birth and who had been given 2 doses of AOTP, developed clinical signs of pertussis infection between 2 and 4 months of age. Pertussis antibody levels immediately prior to infection were not significantly different from those of un infected age-matched cohorts. The percentage of infants afflicted with common childhood illnesses were similar in PEM and WN groups (46 vs. 43.2%) although the former group incurred significantly more illnesses at an earlier age' (s6 months) (p=O.05, chi square). These findings, albeit preliminary given the small numbers of subjects studied, suggest that acellular pertussis vaccine may be incorporated into routine vaccination schedules followed in developing countries with the expectation that adequate antibody responses will be provoked in SGA infants and in infants who develop post-natal PEM and that the incidence of vaccinerelated adverse effects will be no higher than in WN infants. Further and more extensive studies are indicated before the use of acellular pertussis vaccines can be recommended for routine primary vaccination of infants in preference to whole-cell preparations in developing countries.
Optimum timing for vitamin A supplementation in children with diarrhoea.
(2001) Elson, Karin Inga.; Coutsoudis, Anna.
Vitamin A has well recognised benefits for the reduction in severity of diarrhoeal episodes but the impact of therapeutic doses given during diarrhoea on the biochemical and clinical outcomes is less clear. In this study these potential therapeutic benefits were investigated to establish the optimum time for vitamin A supplementation to improve vitamin A status. Establishing the optimum time for vitamin A supplementation during an infectious stage would improve cost effectiveness and clinical benefit. Young children (174) between the ages of 3 and 60 months with severe diarrhoea were randomised in a double - blinded placebo controlled trial into one of 2 groups. The 1 st group received 60 mg of retinol as retinyl palmitate on admission during the acute diarrhoeal stage. The 2nd group received the same dose of vitamin A once symptoms had resolved, usually between 3 - 7 days. At each of these two time points, children not receiving vitamin A were given an identical placebo dose. Baseline (day 0) and day 3 serum samples were collected for vitamin A, retinol binding protein (RBP) and other biochemical markers. At four and eight weeks after discharge both morbidity and weight gain were recorded. The modified dose response test (MRDR) was conducted at the eight - week follow - up to estimate vitamin A liver stores. Initially, most of the children presented with watery diarrhoea and dehydration and were clinically very ill. At day 3 plasma retinol concentrations improved in both groups viz. from 0.57umol/L to 0.97umol/L in the 1st group and from 0.49umol/L to 0.90umol/L in the 2nd group. Similar improvements were found in retinol binding protein viz. 21.28 mg/L to 31.06 mg/L in the 1st group and 17.05 mg/L to 24.80 mg/L in the 2nd group. At 8 weeks there was also no significant difference between the two groups either for serum retinol (0.69umol/L and 0.73umol/L respectively) nor for MRDR ratios (0.036 and 0.049 respectively). The MRDR results at 8 weeks indicated that these children did not have depleted vitamin A liver stores and that the low serum retinol levels seen at baseline were probably due to the acute phase response during an infectious episode. The results of these analyses showed no significant difference between the two treatment groups thus indicating that there was no benefit to giving vitamin A on recovery from an infectious episode instead of on admission, as is currently practised.
Transmission rates of HIV-1 and the mortality rate in high risk infants exposed to HIV, in the PMTCT programme, at the Neonatal Unit, of King Edward VIII Hospital , Durban, South Africa.
(2012) Nair, Nadia.; Adhikari, Miriam.
Introduction. Previous studies have established that infants born to mothers with advanced HIV disease and co-infections are smaller, premature and have rapidly progressive HIV disease and an early death. King Edward VIIIth Hospital, in Durban, admits many sick mothers and manages a large proportion of low birth weight and ill newborns. On discharge and follow-up, the mortality and morbidity of these infants are known to be high and are related to the prematurity. How much is related to being HIV exposed is still uncertain. Aim. To determine the perinatal transmission rate of HIV-1 and mortality at 12 months in HIV exposed infants that were admitted to and discharged from the Neonatal Unit, in Durban, South Africa. Methods. In this observational study, data from the outpatient charts of HIV exposed infants that required specialised neonatal care and subsequent follow up, between the period November 2007 and December 2009, were collected. Perinatal transmission rates and mortality of these infants were compared with maternal and infant risk factors. Results. Data on 463 HIV exposed, predominantly low birth weight infants are presented. The median maternal CD4 count was 309cells/mm3 with 16.8% of mothers commenced on HAART. Maternal co-infection with TB was found in 19.2% of the cohort. Early HIV transmission occurred in 11.5% of infants and was influenced by the type of ARV exposure (None, 20%; single dose NVP, 14.3%; dual therapy, 10.6%; maternal HAART, 8.5%). The dual therapy regimen for 7 days was more protective than that for 28 days (p=0.045). HIV infection was associated with higher risk of neonatal sepsis (RR 1.6; 95% CI, 1.1-2.3; p=0.015). The mortality for the cohort at 12 months was 10%. Maternal HAART was associated with a lower mortality: 2.95% vs.10.2% (RR 3.0; 95% CI, 0.4-20.5). There was a higher mortality rate in those that were low birth weight (RR 4.2; 95% CI, 1.02-18.8; p=0.037); those that were HIV infected (RR 4.8; 95% CI, 1.9-11.6; p=0.002) and those that were breastfeeding compared to formula feeding (RR 2.7; 95% CI, 1.1-6.8; p=0.038). Discussion. Rates of HIV transmission within the PMTCT programme were similar to that reported by the Department of Health. Early maternal ARVs for PMTCT prophylaxis, prevents HIV transmission. The coverage of maternal HAART was sub-optimal. Breastfeeding was associated with a higher HIV transmission rate and was most likely associated with non-exclusive breastfeeding during neonatal admission. Recommendations. Maternal HAART or ARV prophylaxis should be commenced early in the pregnancy for the best benefits. Meticulous attention should be paid to the feeding practices of high risk HIV exposed infants admitted for specialised neonatal care.
Neutralizing antibody responses and viral evolution in a longitudinal cohort of HIV subtype C infected antiretroviral-naïve individuals.
(2011) Archary, Derseree.; Coovadia, Hoosen Mahomed.; Ndung'u, Peter Thumbi.
Background: HIV-1 envelope (Env) diversity is arguably the most significant challenge for the development of an efficacious vaccine. An ideal vaccine would elicit the production of broadly neutralizing antibodies (nAb), capable of retaining potent activity against a diverse panel of viral isolates. The evolutionary forces that shape the diversity of envelope and ensuing nAb responses are incompletely understood in HIV-1 subtype C infection, the dominant subtype globally. Therefore there is an urgent need to define the patterns of envelope diversity, determine the correlates of immune protection and to discover subtype C immunogens in order to develop a globally relevant vaccine. Methods: We applied the single genome sequencing strategy to study plasma derived viruses from four slow progressors and four progressors over a median of 21 months between study entry and study exit. The participants‘ samples were from the Sinikithemba cohort of antiretroviral therapy-naïve chronically infected individuals and were termed slow progressors or progressors based on CD4 T-cell counts and viral loads over two years. We analyzed env sequence diversity, divergence patterns and envelope characteristics across the entire HIV-1 subtype C gp160. We studied the evolution of autologous nAb (AnAb) and heterologous nAb responses in order to test the hypothesis that slow disease progression is associated with more potent autologous or heterologous nAb responses. Furthermore, genotypic env characteristics were correlated to potency of neutralization in order to understand possible differences in nAb responses with divergent rates of disease progression and to describe genotypic differences associated with differential nAb potencies. In addition, the binding affinities of HIV-specific immunoglobulins (IgGs) and the affinities of the IgGs to various Fcγ receptors (both activating- FcγRI, FcγRIIa, FcγRIIIa; inhibitory- FcγRIIb) were assessed. These binding affinities were used as a surrogate for the recruitment of effector functions of cells of the innate immune system e.g. macrophages or natural killer cells to initiate antibody-dependent cell-mediated cytotoxicity (ADCC) or antibody dependent cell-mediated viral inhibition (ADCVI) and these were correlated to markers of disease progression namely CD4 T-cell counts and viral loads. Results: Intra-patient diversity was higher in slow progressors for regions C2 (p=0.0006), V3 (p=0.01) and C3 (p=0.005) compared to progressors. Consistent with this finding, slow progressors also had significantly increased amino acid length in V1-V4 with fewer potential N-linked glycosylation sites (PNGs) compared to progressors (p=0.009 and p=0.02 respectively). Similarly, in progressors, the gp41 region was significantly longer and had significantly fewer PNGs compared to slow progressors (p=0.02 for both parameters). Positive selection was prominent in regions V1, C3, V4, C4 and gp41 in slow progressors, whereas in progressors, it was prominent in gp41. Signature consensus sequence differences between the groups occurred mainly in gp41. Neutralizing antibodies (nAb) evolved over time in progressors, as evidenced by significantly higher nAb IC50 titers to baseline (study entry) viruses when tested against study exit time-point plasma compared to contemporaneous responses (p=0.003). In contrast, slow progressors‘ nAb titers did not differ significantly between study entry and study exit time points. nAb IC50 titers significantly correlated with amino acid lengths for C3-V5 (p=0.03) and V1-V5 (p=0.04) for slow progressors and V1-V2 for progressors (p=0.04). Slow progressors and progressors displayed preferential heterologous activity against the subtype C panel. There were no significant differences in breadth of responses between the groups for either subtype A or C. Neutralization breadth and titers to subtype B reference strains however, was significantly higher in progressors compared to slow progressors (both p<0.03) with increasing nAb breadth from study entry to study exit in progressors. Progressors had cross-reactive neutralizing antibodies that targeted V2 and V3. Binding affinities of non-neutralizing antibodies to HIV-specific gp120, gp41 and p24 and to activating and inhibitory Fcγ receptors (FcγRs) were similar in both groups. However, in slow progressors, CD4 T-cell counts correlated inversely with antibody binding affinity for the activating FcγRIIa (p=0.005). Conclusions: These data suggest that separate regions of Env are under differential selective forces, and the heterogeneity of env diversity and evolution differ with HIV-1 disease course. Single genome sequence analysis of circulating viruses in slow progressors and progressors indicate that diversity, length polymorphisms, sites under positive selection pressure, and PNGs consistently map to specific regions in Env. Cross-reactive neutralizing antibodies targeting epitopes in V2 and V3 indicate that nAb breadth may be dictated by a limited number of target Env epitopes. Certain key N-linked glycosylation sites were shown to be crucial for antibody neutralization. The potencies of autologous nAbs were directly affected by the amino acid lengths in certain regions of Env gp160 and by the numbers of PNGs. Target vaccine immunogens may have to be given over long periods of time and may have to include multiple subtype immunogens to elicit the production of potent, broad cross neutralizing antibodies with high binding affinity. Overall, the data suggest that neither nAbs nor non-neutralizing antibodies could be directly associated with disease attenuation in this cohort of chronically infected individuals. However, continuous evolution of nAbs was a potential marker of HIV-1 disease progression. Further studies on larger cohorts to identify people with potent nAbs and to identify specific targets of these antibodies are needed. Furthermore studies of non-neutralizing antibodies in HIV-1 infection using functional assays will be required in order to determine their role in HIV-1 pathogenesis.
Effect of infant feeding mode and maternal nutritional supplementation on the nutrition and health of HIV positive mothers and their infants.
(2012) Kindra, Gurpreet.; Coutsoudis, Anna.
Background: Breastfeeding is known to have benefits both for maternal and child health. Some questions around the benefits and risks of breastfeeding in the presence of HIV infection still remain unclear. Aims: To study the effects of infant feeding mode by HIV-positive mothers, on maternal and child health. In addition, to assess the effect of nutritional supplementation to HIV-positive lactating mothers on nutritional and health status of mothers and their infants and on the quality of breastmilk. Methods: The study had 2 components; a prospective study to examine the impact of infant feeding mode on nutritional and health indices in mothers and their infants and within it a nested randomized controlled clinical trial to study the impact of a daily 50 g soya/peanut based supplement during breastfeeding on the above parameters. The measurements included anthropometry; body composition indicators (using both deuterium dilution and BIA); haematology and biochemical markers; as well as incidence rates of opportunistic infections and clinical disease progression. Breastmilk was analysed for both macro and micronutrients. Cervical screening was offered to all the women. Results: AFASS criteria were fulfilled by 38.7% of the formula feeding mothers. No significant differences between the formula feeding and breastfeeding groups in terms of haematological, immunological and body composition changes were seen. Breastfeeding mothers had significantly lower events with high depression scores (p=0.043). Longer duration of breastfeeding was observed to be significantly associated with a mean increase in CD4 count (74 cells/μL) and better health outcomes. The supplement made no significant impact on any maternal or child outcomes except for a limited effect on mothers with low BMI, where it was significantly associated with preventing loss of lean body mass (p=0.026). Breastfeeding infants had a significantly lower risk of diarrhoea and hospitalisation at 3 months (p=0.006 and 0.014 respectively). Both breastfeeding and longer duration of breastfeeding was significantly associated with better development scores and growth parameters. Supplementation made no impact on breastmilk composition. Of the 86 mothers who agreed for cervical screening, 27.6% had human papilloma virus infection. Conclusions: Breastfeeding is not harmful to the mother despite the presence of HIV infection. On the contrary we observed both breastfeeding and longer breastfeeding duration to be associated with better maternal and child outcomes. Mothers are still choosing formula feeding inappropriately presumably because of the availability of free formula and/or sub-optimal counseling. The new (2010) local PMTCT guidelines based on WHO recommendations should reverse this. Food insecurity was prevalent amongst 32% of our study population, highlighting the need to include sustainable and empowering solutions to encounter this problem. Less sustainable solutions such as nutritional supplementation should be targeted to the malnourished and in emergency situations.
Is IMCI an effective mechanism for delivery of child survival interventions in a high HIV prevalence setting? : a study to determine the effectiveness of the Intergrated Management of Childhood Illness (IMCI) strategy in management of sick children in routine practise in primary health care clinics in South Africa
(2012) Horwood, Christiane.; Rollins, Nigel C.; Adhikari, Miriam.
Introduction: Integrated management of childhood illness (IMCI) is a child survival strategy that has been adopted in South Africa (SA) as the standard of care for managing sick children in the primary health care setting. IMCI includes guidelines for management of paediatric HIV. This study aimed to investigate effectiveness of IMCI as a vehicle to deliver essential child survival interventions, particularly HIV interventions, in routine practise in a high HIV prevalence setting, and to investigate barriers and enabling factors for IMCI implementation. Methods: The study was conducted in Limpopo and KwaZulu-Natal provinces, SA. In the qualitative component, focus group discussions were conducted with IMCI trained health workers and carers of children under 5 years, to explore experiences of IMCI implementation, particularly the HIV component, from the perspective of both target groups. A comparative survey was then conducted. Randomly selected IMCI trained nurses were observed for up to 20 consultations with sick children presenting consecutively to the facility, and their findings compared to those of an IMCI expert who subsequently assessed the child. Observed children were tested for HIV. Results: IMCI trained nurses found IMCI training informative and empowering, and there was agreement among nurses that their skills in managing sick children improved after training. Barriers to IMCI implementation included increased time required for IMCI consultations and lack of support from colleagues. IMCI trained nurses expressed reluctance to implement the HIV component of IMCI, believing it to be unnecessary, unacceptable to mothers and that they lacked the skills to implement HIV care. In total, 77 IMCI trained nurses were observed for a total of 1357 consultations between May 2006 and January 2007; nurses were observed for a mean of 17.7 consultations. Components of the IMCI assessment were frequently omitted; 14/77(18%) nurses asked about all main symptoms in every child. IMCI classifications were often incorrect; 52/112 (46.4%) children with a general danger sign were correctly classified. The HIV component was poorly implemented, 342/1357 (25.2%) children were correctly classified for HIV, although the HIV algorithm performed well when implemented by IMCI experts. Conclusion: IMCI implementation is fragmented and incomplete. Interventions are urgently needed to achieve and maintain high quality health worker performance in implementing IMCI.
Coreceptor utilization and primary cell tropism by HIV-1 subtype C strains.
(2010) Singh, Ashika.; Ndung'u, Peter Thumbi.
Human immunodeficiency virus type 1 (HIV-1) isolates can be differentiated based on their ability to use particular coreceptors – R5 viruses use CCR5, X4 viruses use CXCR4 and R5X4 (dual tropic) viruses use both CCR5 and CXCR4. It is widely reported that HIV-1 subtype C (HIV-1C) has a unique viral coreceptor evolution pattern in that a complete switch from the predominant CCR5 (R5) to CXCR4 (X4) phenotype is less common for this subtype compared to other subtypes. However, dual tropic HIV-1C isolates have occasionally been described. Furthermore, it has been reported that certain highly active antiretroviral drugs (HAART) may select for X4 viral variants. Therefore, this thesis study was undertaken to better understand the functional and genotypic characteristics of dual tropic HIV-1C isolates, and to characterize drug resistance and coreceptor usage patterns in HAART-naïve versus HAART-failing HIV-1C infected patients. Thirty-five functional HIV-1 env clones derived from seven dual tropic HIV-1C strains were generated and their coreceptor usage characterized in transformed cell lines. All 35 env clones efficiently infected transformed cells expressing CXCR4. Twenty of 35 clones (57%) also utilized the CCR5 receptor. No R5-only clones were detected. Functional coreceptor usage data was correlated to env gene sequence data. The ability of the HIV-1C env clones to facilitate infection of primary lymphocytes and monocyte-derived macrophages was next investigated. The majority of clones characterized as X4 or R5X4 on cell lines used either CXCR4 alone or CXCR4 and CCR5, respectively, in primary cells. A few viruses displayed comparable CCR5 and CXCR4 usage and clones from one virus preferred CCR5 usage in macrophages. Thus in a few cases coreceptor phenotyping in transformed cell lines does not predict usage in primary cells. Genetic determinants for coreceptor usage in primary cells require further investigation. Finally the patterns of drug resistance mutations were studied and coreceptor usage among 45 HAART-naïve and 45 HAART-failing HIV-1C infected patients analyzed. Ninety-five percent of HAART-failing patients had viruses with at least one drug resistance mutation. Thymidine analog resistance mutations (TAMs) were present in 55% of patients. HAART-failing patients had significantly higher prevalence (59%) of X4/R5X4-utilizing viruses compared to HAART-naïve patients (30%) (p<0.02) using the Trofile Co-receptor Tropism Assay while 41% of HAART-failing patients used CCR5 and 70% of HAART-naïve patients used CCR5. Functional results correlated with predictive algorithm methods. This study enhances our understanding of HIV-1 pathogenesis and the results have important implications for the use of coreceptor antagonists for the clinical management of HIV-1C infection.
The role of HLA-C restricted CD8+T cell responses in the control of HIV replication.
(2010) Mkhwanazi, Nompumelelo Prudence.; Ndung'u, Peter Thumbi.
Certain HLA-B-restricted CD8+ T cell responses are associated with control of viremia whereas HLA-Cw* restricted responses, including Gag epitopes are associated with high viremia. To better understand the role of HLA-Cw* restricted epitopes in viral control, HLA-Cw* restricted epitopes were optimally defined. Seventy eight study subjects from a cohort of 451 chronically infected participants had HLA-Cw* restricted CD8+ T cells responses as quantified by intracellular cytokine staining assessing IFN-γ secretion. Fine mapping and HLA restriction of the optimally defined HLA-Cw* restricted epitopes were performed using ELISPOT assay. Functional avidity of responses was assessed by peptide dilution in an ELISPOT assay. Two novel HLA-Cw* restricted epitopes Cw*04- TF10 (in reverse transcriptase) and Cw*08-RM9 (in gp120) were optimally defined. A previously described epitope, Cw*07- KY11 (Nef) was the most frequently targeted epitope in this cohort (30/78) and has high functional avidity compared to other HLA-Cw restricted CD8+ T cell responses. The polyfunctionality of HLA-B*57/5801-restricted Gag-specific HIV-1 CD8+ T cell responses and HLA-Cw*07-KY11 restricted CD8+ T cell responses within the same study subject was determined. Polyfunctionality of CD8+ T cell responses to HLAB* 57/5801 and HLA-Cw*07 restricted epitopes were determined in nine study subjects assessing IFN-γ, TNF-α, IL-2, MIP-1β, and CD107a by multicolour flow cytometry. Additionally gag and nef genes were sequenced from plasma. HLA-B*57/5801-restricted IFN-γ-producing CD8+ T cell responses were of lower magnitude than HLA-Cw*07 responses (p=0.0012) for the nine subjects. The majority of responses were monofunctional (75%), irrespective of HLA restriction. HLA-B*57/5801 and HLACw* 07 restricted CD8+ T cells did not differ significantly in polyfunctionality (p=0.84). Possession of ≥3 functions correlated positively with CD4+ T cell counts (r=0.85; p=0.006). The percentages of monofunctional CD8+ T cells inversely correlated with CD4+ T cell counts (r=-0.79; p=0.05). There was no correlation between polyfunctionality and viral load and sequence variation within targeted epitopes did not impact polyfunctionality. These results suggest that polyfunctionality of HIV-1-specific CD8+ T cells is associated with disease progression independent of restricting HLA alleles, and that loss of these polyfunctional cells correlates with increased in the frequency of monofunctional virus-specific CD8+ T cells. In addition, sequence variation does not appear to significantly impact CD8+ T cell polyfunctionality in chronic HIV infection.
The association of early neonatal feeding on clinical outcomes and cytotoxic T lymphocyte (CTL) responses in HIV exposed low birth weight infants.
(2011) Dassaye, Reshmi.; Coutsoudis, Anna.; Sunpath, Henry.
BACKGROUND Sub-saharan Africa remains to date at the forefront of the HIV/AIDS epidemic. Despite breastfeeding being a significant mode of postnatal HIV transmission it remains the main nutritional source and pillar of child survival for the majority of infants born in Africa. It is therefore, not surprising that considerable research has centred on making breastfeeding safer in terms of HIV transmission. The flash heat treatment method (HTEBM) provides a unique opportunity to safely breastfeed infants but prevent mother-to-child transmission of HIV. Cytotoxic T lymphocyte (CTL) responses have been well documented in HIVinfected adults and children. However, there is a lack of literature on CTL responses in HIV exposed low birth weight infants. This pilot study attempted to examine the association of early neonatal feeding on the clinical outcomes and CTL responses in HIV exposed low birth weight infants. METHODS Seventy-seven patients that fulfilled inclusion and exclusion criteria were enrolled. The clinical outcomes of these patients were evaluated over a 9 month period. Fifty-five of these patients were also investigated for cytotoxic T lymphocyte (CTL) responses by means of the IFNγ ELISpot (megamatrix and confirmation) assays at the 6 weeks, 3, 6,and 9 months follow-up. RESULTS Two HIV-1 infected infants generated a CTL response at a single time point using the ELISPOT matrix screening assay. These responses could not be confirmed and were undetectable at any of the consecutive visits. At the time of detection of responses the infants were fed unheated breastmilk. HIV-1 exposed uninfected infants were unable to elicit a HIV-1-specific CTL response irrespective of feed. With regards to clinical outcomes, infants born o HIV infected mothers with a CD4 count < 500cells/μl were 2x more likely to acquire other infections at birth compared to those infants born to HIV infected mothers with a CD4 count >500cells/μl. Also, infants born to HIV infected mothers with advanced disease (CD4 count 0-200 cells/μl) had a lower birth weight compared to infants born to HIV-1 infected mothers with a CD4 count > 350 cells/μl. We also investigated the feasibility of the flash heat treatment method at birth. While inhospital, 38 HIV-1 infected women fed their infants HTEBM after receiving counseling and support from the nursing staff at the King Edward VIII hospital. The numbers decreased rapidly post hospital discharge, mainly due to mixed feeding. DISCUSSION In conclusion we have shown that it is feasible for HIV infected mothers to heat treat their expressed breastmilk during hospital admission. Furthermore, we were able to demonstrate in this small cohort of patients that the clinical outcomes and growth parameters of infants fed HTEBM were similar to that of infants fed either formula or unheated breastmilk. We were unable to demonstrate HIV-specific responses in the infected infants or the uninfected infants who had been exposed to heat inactivated virus in HTEBM. Our findings indicate that this pilot study was limited in its ability to detect CTL responses in HIV exposed low birth weight infants and further studies are warranted.
Cellular immunity, immune activation and regulation in HIV-1 infected mother-child pairs : what are the determinants of protective immunity.
(2011) Moodley-Govender, Eshia S.; Ndung'u, Peter Thumbi.; Addo, Marylyn.; Goulder, Philip Jeremy Renshaw.
Background: Prevention of Mother-to-child transmission (PMTCT) of human immunodeficiency virus (HIV) remains a significant challenge in resource-poor settings despite the advances in antiretroviral (ARV) treatment. HIV-1 infected individuals are able to achieve viral control naturally, however the underlying mechanisms of immunological control in children remains poorly understood. This study was conducted from 2006 to 2010 to investigate correlates of immune control in HIV-1 clade C infected mother-child pairs in the absence of ARVs. Genotypic and phenotypic viral characteristics, cellular immune responses to HIV-1 and host genetics were characterized and correlated with clinical markers of disease progression. Materials and Methods: To achieve the objectives of the study, three cohorts of mother-child pairs were investigated. The first cohort included 60 untreated mother-child pairs and a further ten uninfected children as controls. The second cohort comprised of ARV treated pairs (n=60). The third cohort consisted of 374 mothers and 374 children (infected, exposed uninfected, HIV negative). Plasma viral loads and absolute CD4+ T cell counts were routinely performed in all three cohorts. HIV-specific CD8+ T cell responses were analyzed by interferon gamma (IFN-γ) enzyme linked immunosorbent spot (ELISpot) assays. Viral replicative fitness was assessed using a green fluorescent protein reporter cell line (GFP).Multi-parameter flowcytometry allowed for the investigation of T cell regulation, exhaustion and activation using CD127/CD25, TIM-3/PD-1 and HLA-DR/CD38 markers respectively. IL-10 promoter single nucleotide polymorphisms (SNPs) at positions -592 and -1082 were determined by TaqMan allelic discrimination assays. Plasma IL-10 levels were measured using a luminex assay. Results: To describe the CTL responses elicited to various regions of the HIV proteome in HIV-infected treatment naïve children. Sixty children under one year of age in the untreated cohort were analyzed for CTL responses spanning the HIV genome, for which only 30 had detectable responses. There was no significant difference in viral load between respondersand non-responders (p=0.2799). The responders predominantly targeted Nef (49%), Gag (17%) and Env (14%) regions. Markers of T cell exhaustion and regulation and theirrelationship to markers of disease progression, were next investigated as these parameters may explain the inability of T cells to effectively control HIV infection. T cell phenotyping compared treated, untreated and uninfected subgroups. In infected children, CD8+ T cells were significantly higher for both the inhibitory marker TIM-3 (p=0.001) and exhaustion marker PD-1 (p=0.0001) compared to uninfected children. Median expression of TIM-3 was higher on CD8+ T cells (46%) compared to CD4+ T cells (20%). TIM-3 and PD-1 expression on T cells were maintained at high levels over time. The frequency of absolute Tregs (p=0.0225) were found to be significantly higher in untreated compared to treated children. HLA-DR+CD38+ on CD8+ T cells were significantly up-regulated in untreated children compared to treated (p=0.002) and uninfected children (p=0.0177). HLA-DR+CD38+ was also significantly higher in children less than 6 months compared to older children on CD4+ (p=0.0437) and CD8+ T cells (p=0.00276). Interestingly, we observed a significant negative correlation between magnitude of CTL response and CD25+CD127- (p=0.0202; r=-0.7333) as well as HLA-DR+CD38+ (p=0.0408; r=-0.5516) on CD8+ T cells. IL-10 is an important immunoregulatory cytokine that has been shown to affect the outcome of chronic viral infections. IL-10 polymorphisms have previously been associated with IL-10 levels and HIV-1 outcomes in adults. Polymorphisms associated with different levels of IL-10 production and their relationship with transmission, markers of disease progression and immune responses were next investigated in this mother-child HIV transmission setting. Genetic analysis of IL-10 in cohort three revealed that HIV-1 acquisition was not associated with either IL10 -592 (AA/CA vs CC) or IL10 -1082 (AA/AG vs GG) single nucleotide polymorphisms (SNPSs). There was a significant association between IL10 -1082 and HIV-1 transmission (p=0.0012). No correlation was observed between IL10 -592 (p=0.4279) or IL10 -1082 SNPs (p=0.6361) and mortality rates in children. IL10 -592C was associated with an elevated magnitude of IFN-γ CD8+ T cell response compared to IL10 -529A (p=0.0071). We found a significant positive correlation between IL-10 plasma levels and viral loads (p=0.0068; r=0.4759) and the ages of the children (p=0.0312; r=0.1737). Conclusion: CD8+ T cell responses and viral fitness did not explain differences in disease progression in selected HIV-1 untreated clade C transmission pairs. T cell activation and regulatory markers influence CTL immune responses resulting in poor clinical outcome. IL10 -1082 polymorphisms may be used as a predictor of HIV-1 transmission. The association between increased IL-10 plasma levels and high viral loads suggest that IL-10 contributes to immune dysfunction in paediatric HIV-1 infection. This study has extended our understanding of immunological and genetic correlates of mother-to-child transmission and disease outcome in ARV naïve (naturally controlling) and HIV treated infected children.

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