Etiology of diarrhea in children younger than 5 years of age admitted in a rural hospital of southern Mozambique.

Diarrhea is one of the main causes of morbidity and mortality among children in sub-Saharan Africa and one of the main causes of hospital admissions in rural areas. Stool samples were collected from 529 children admitted with diarrhea to the Manhiça District Hospital (September 2000 to September 2001) and processed to detect bacterial enteropathogens, parasites, and virus. Diarrheagenic Escherichia coli, isolated from 120 samples (22.6%; enteroaggregative [corrected] [9.6%], enterotoxigenic [6.8%], enteropathogenic [corrected] [4.3%], and verotoxigenic [1.9%]) was the most frequently isolated pathogen, followed by Ascaris lumbricoides (9.3%). Others detected included Salmonella spp. and Giardia lamblia (2.5% each) and Campylobacter spp. (1.7%). A. lumbricoides (92% versus 8%; P<0.001) and Strongyloides stercolaris (100% versus 0%; P=0.008) were most frequently isolated in children older than 12 months of age. Resistance to trimethoprim-sulphametoxazole and ampicillin was high. Etiologic data on diarrheal diseases and susceptibility patterns of diarrheal pathogens are important tools for clinical management and control strategic planning.

Duration of protection with RTS,S/AS02A malaria vaccine in prevention of Plasmodium falciparum disease in Mozambican children: single-blind extended follow-up of a randomised controlled trial.

BACKGROUND: RTS,S/AS02A is a pre-erythrocytic stage malaria vaccine that provides partial protection against infection in malaria-naive adult volunteers and hyperimmune adults. A previous report showed that this vaccine reduced risk of clinical malaria, delayed time to new infection, and reduced episodes of severe malaria over 6 months in African children. An important remaining issue is the durability of protection against clinical disease in these children. METHODS: We did a randomised, controlled, phase IIb trial of RTS,S/AS02A given at 0, 1, and 2 months in 2022 Mozambican children aged 1-4 years. We previously determined vaccine efficacy (VE) against clinical malaria in a double-blind phase that included study months 2.5-8.5 (VE(2.5-8.5)). We now report VE in a single-blind phase up to month 21 (VE(8.5-21)). The primary endpoint was time to first or only clinical episode of Plasmodium falciparum malaria (axillary temperature 37.5 degrees C and P falciparum asexual parasitaemia >2500 per microL) detected through a passive case detection system. We also determined VE for other case definitions and for episodes of severe malaria. This study is registered with the ClinicalTrials.gov identifier NCT00197041. FINDINGS: During the single-blind phase, VE(8.5-21) was 28.9% (95% CI 8.4-44.8; p=0.008). At month 21, prevalence of P falciparum infection was 29% lower in the RTS,S/AS02A group than in the control (p=0.017). Considering the entire study period, VE(2.5-21) was 35.3% (95% CI 21.6-46.6; p<0.0001) and VE(2.5-21) for severe malaria was 48.6% (95% CI 12.3-71.0; p=0.02). INTERPRETATION: These results show that RTS,S/AS02A confers partial protection in African children aged 1-4 years living in rural endemic areas against a range of clinical disease caused by P falciparum for at least 18 months, and confirm the potential of malaria vaccines to become credible control tools for public-health use.

Efficacy of the RTS,S/AS02A vaccine against Plasmodium falciparum infection and disease in young African children: randomised controlled trial.

BACKGROUND: Development of an effective malaria vaccine could greatly contribute to disease control. RTS,S/AS02A is a pre-erythrocytic vaccine candidate based on Plasmodium falciparum circumsporozoite surface antigen. We aimed to assess vaccine efficacy, immunogenicity, and safety in young African children. METHODS: We did a double-blind, phase IIb, randomised controlled trial in Mozambique in 2022 children aged 1-4 years. The study included two cohorts of children living in two separate areas which underwent different follow-up schemes. Participants were randomly allocated three doses of either RTS,S/AS02A candidate malaria vaccine or control vaccines. The primary endpoint, determined in cohort 1 (n=1605), was time to first clinical episode of P falciparum malaria (axillary temperature > or =37.5 degrees C and P falciparum asexual parasitaemia >2500 per microL) over a 6-month surveillance period. Efficacy for prevention of new infections was determined in cohort 2 (n=417). Analysis was per protocol. FINDINGS: 115 children in cohort 1 and 50 in cohort 2 did not receive all three doses and were excluded from the per-protocol analysis. Vaccine efficacy for the first clinical episodes was 29.9% (95% CI 11.0-44.8; p=0.004). At the end of the 6-month observation period, prevalence of P falciparum infection was 37% lower in the RTS,S/AS02A group compared with the control group (11.9% vs 18.9%; p=0.0003). Vaccine efficacy for severe malaria was 57.7% (95% CI 16.2-80.6; p=0.019). In cohort 2, vaccine efficacy for extending time to first infection was 45.0% (31.4-55.9; p<0.0001). INTERPRETATION: The RTS,S/AS02A vaccine was safe, well tolerated, and immunogenic. Our results show development of an effective vaccine against malaria is feasible.

Characterization of an integron carrying a new class D beta-lactamase (OXA-37) in Acinetobacter baumannii.

Integrons from a clinical strain of Acinetobacter baumannii highly resistant to beta-lactams have been analyzed. The largest (2.2 kb) contained three gene cassettes: an aacA4, an open reading frame of 417 nucleotides, and an OXA-type encoding gene. The oxa gene nucleotide sequence differed from that of the oxa-20 in 2 bp, one of the mutations being silent. The nonsilent mutationgenerated a substitution of glutamic acid for aspartic acid. The new OXA has been named OXA-37.

Detection of dihydrofolate reductase genes by PCR and RFLP.

The presence of plasmid-encoded trimethoprim resistant dfr genes is the most common mechanism responsible for the acquisition of trimethoprim resistance. The usual method to detect the presence of these genes is hybridization with specific probes. We describe an alternative, faster and easier method, based on PCR amplification and RFLP analysis, to discriminate up to sixteen different dfr genes.

High prevalence of nalidixic acid resistant, ciprofloxacin susceptible phenotype among clinical isolates of Escherichia coli and other Enterobacteriaceae.

Therapeutic failure of infections during their treatment with quinolones has been often described. This may be due to the development of resistance during treatment of an infecting strain which already had diminished susceptibility to quinolones, even though the initial MIC did not exceed the breakpoint. In this study the prevalence of the nalidixic acid resistant, ciprofloxacin susceptible phenotype among Enterobacteriaceae was analyzed. The results showed that 113 out of 151 (74.83%) strains of the Enterobacteriaceae with diminished susceptibility to ciprofloxacin (MICs from 0.06 to 1 microg/ml) were resistant to nalidixic acid (MICs > 32 microg/ml). The Escherichia coli strains presenting this phenotype already have a mutation in the amino acid codon Ser-83 of the gyrA gene, so that the possibility of developing a second mechanism of resistance during treatment is very high.

Analysis of the clonal relationship among clinical isolates of Salmonella enterica serovar Infantis by different typing methods.

Salmonella Infantis has been the second most common serovar in Argentina in the last two years, being isolated mostly from paediatric hospitalised patients. In order to determine the clonal relationship among Salmonella Infantis strains, we examined 15 isolates from paediatric patient faeces in Argentina (12 geographically related and 3 geographically non-related) by using antimicrobial susceptibility, plasmid profiling, repetitive extragenic palindromic (REP) PCR, enterobacterial repetitive intergenic consensus (ERIC) PCR, and low-frequency restriction analysis of chromosomal DNA by pulsed field gel electrophoresis (PFGE). Four Spanish strains were included as controls of clonal diversity in molecular techniques. Antibiotype and plasmid profile was not useful as epidemiological tools. PFGE and REP-PCR were able to discriminate between Argentinean and Spanish isolates of Salmonella Infantis allowing to detect genetically related strains in three different cities. This finding indicates that a possible spread of a clone of this serovar in the North-eastern Region of Argentina has taken place in 1998.

Evaluation of two formulations of adjuvanted RTS, S malaria vaccine in children aged 3 to 5 years living in a malaria-endemic region of Mozambique: a Phase I/IIb randomized double-blind bridging trial.

BACKGROUND: Previous trials of the RTS, S malaria candidate vaccine have shown that this vaccine is safe, tolerated and immunogenic. The development plan for this vaccine aims at administering it in the first year of life through the Expanded Program on Immunization (EPI). The objective was to evaluate the safety and reactogenicity of RTS, S/AS02D (0.5 ml dose), a pediatric formulation of GlaxoSmithKline Biologicals' current malaria candidate vaccine RTS, S/AS02A (0.25 ml dose). A 0.5 ml dose of AS02D is composed of the same active ingredients in the same quantities as in a 0.25 ml dose of AS02A and has been developed to be easily introduced into routine EPI practices. METHODS: We performed a phase I/IIb randomized double-blind bridging study in a malaria-endemic region of Mozambique, to compare the safety and immunogenicity of both candidate vaccines with the aim of replacing RTS, S/AS02A with RTS, S/AS02D as the candidate pediatric vaccine. 200 Mozambican children aged 3 to 5 years were randomized 1:1 to receive one of the 2 vaccines according to a 0, 1, 2 month schedule. RESULTS: Both vaccines were safe and had similar reactogenicity profiles. All subjects with paired pre and post-vaccination samples showed a vaccine response with respect to anti-circumsporozoite (CS) antibodies irrespective of initial anti-CS serostatus. Geometric mean titers (GMTs) were 191 EU/ml (95% CI 150-242) in recipients of RTS, S/AS02D compared to 180 EU/ml (95% CI 146-221) in recipients of RTS, S/AS02A. For the anti-hepatitis B surface antigen (HBsAg), all subjects were seroprotected at day 90, and the GMTs were 23,978 mIU/ml (95% CI 17,896-32,127) in RTS, S/AS02D recipients and 17,410 mIU/ml (95% CI 13,322-22,752) in RTS, S/AS02A recipients. There was a decrease in anti-CS GMTs between months 3 and 14 in both groups (191 vs 22 EU/mL in RTS, S/AS02D group and 180 vs 29 EU/mL in RTS, S/AS02A group). CONCLUSION: Our data show that the RTS, S/AS02D is safe, well tolerated, and demonstrates non-inferiority (defined as upper limit of the 95% confidence interval of the anti-CS GMT ratio of RTS, S/AS02A to RTS, S/AS02D below 3.0) of the antibody responses to circumsporozoite and HBsAg induced by the RTS, S/AS02D as compared to the RTS, S/AS02A.

Haematological and biochemical indices in young African children: in search of reference intervals.

INTRODUCTION: The reference intervals of haematological and biochemical indices currently used in Africa are derived from data collected from populations living in industrialized countries. Few studies have been performed in Africa questioning the validity of these values when applied to local African populations. OBJECTIVE: To provide reference intervals of haematological [haemoglobin (Hb), white blood cells (WBC), haematocrit (Htc) and platelets] and biochemical indices (ALT, creatinine and bilirubin) for children aged 1-4 from a rural area of southern Mozambique. METHODS: Reference intervals were developed using the 2.5 and 97.5 centiles. Partition tests were performed to evaluate age and gender differences. Quantile regression models were estimated for those variables in which age partition was recommended. Deviances from linearity in the estimated models were evaluated using fractional polynomials of first or second degree. Agreement to classify normality, using the estimated reference intervals or values in use in a western paediatric hospital, was made using the kappa statistic. RESULTS: Reference intervals for Hb, WBC, Htc, platelets, ALT and creatinine show significant differences by age. Gender differences were observed for creatinine values, while for bilirubin there were no significant differences for age or gender. Estimated Hb and Htc reference intervals in African children were lower than the accepted western ones, while ALT values were higher in the former. Agreement between normal classification, using the estimated intervals or the western values, was from slight to fair. CONCLUSIONS: Reference intervals of haematological and biochemical indices based on results from western individuals of the same age are not in agreement with the estimated values for African children. These observed values should not be used as a rule to define normality, but are the ones usually found in this population where anaemia, malaria and high mortality rates are also common.

Relationship between haemoglobin and haematocrit in the definition of anaemia.

INTRODUCTION: Anaemia is the most frequent haematological disorder in childhood. The notion that defines naemia does not change throughout life, although parameters used for its evaluation show significant variations during childhood. Haematocrit (Hct) (%) is usually defined as three times the value of haemoglobin (Hgb) (g/dl), while the clinical definition of anaemia is related to either an abnormal Hct or Hgb value. OBJECTIVE: To evaluate the agreement between Hgb and Hct values in the definition of anaemia, the relationship between these two parameters and their age-dependence. METHODS: The Hct and Hgb paired values from children aged 2-18 months from Ifakara (Tanzania) and children aged 1-4 years from Manhiça (Mozambique) were analysed. Haematological determinations of the Manhiça samples were done using a KX-21N cell counter (Kobe, Japan) and Ifakara samples were analysed in a semiautomatic cell counter (Sysmex F800 microcell counter, TOA Medical Electronics, Kobe, Japan). The kappa-statistic was used to calculate the agreement between anaemia definitions in each group. Crude and multivariate relationship between Hct and Hgb levels were analysed by linear regression model estimation. The age-dependence of the crude ratio (Hct/Hgb) was analysed using linear regression models and fractional polynomials. RESULTS: The prevalences of mild and moderate anaemia as defined by Hgb levels in the Manhiça group were 61% and 6%, respectively, and 41% and 2% by Hct. In the Ifakara group these were 74% and 10%, respectively, by Hgb and 42% and 3% by Hct, respectively. Agreement between mild and moderate anaemia definitions made up from Hgb or from Hct levels were from fair to moderate. Hct levels decreased with age for high Hgb levels, whereas they increased for low Hgb levels. The classification of cases is improved when higher age-related cut-off values for Hct are used. The crude relationship between Hct and Hgb levels was significantly different from 3, and this was modified by age. The evaluation of the age-dependence ratio (Hct/Hgb) showed a non-linear relationship with an asymptotic trend to 3. CONCLUSIONS: Measurement of haematocrit count is easy and can be performed in most rural health care centres. However, the corresponding Hgb levels cannot be derived with an acceptable accuracy using the value 3 as a conversion factor. Furthermore, the commonly assumed 'equivalent' cut-off points for anaemia definitions need to be re-evaluated.

[Intercontinental spread of a trimethoprim-resistant strain of Shigella flexneri]. / Dispersión intercontinental de una cepa de Shigella flexneri resistente a trimetoprima.

INTRODUCTION: In this study we characterize a trimethoprim-resistant strain of Shigella flexneri recovered from stool samples of an international traveler after a trip to Kenya, and analyze its epidemiological relationship with a set of strains having similar characteristics from Tanzania. METHODS: Clonal relationships were studied by three techniques - plasmid profile, repetitive-element (REP)-PCR and pulse-field gel electrophoresis (PFGE). The presence of type 1 integrons was studied by PCR and sequencing. The location and transferability of the detected integron was analyzed by conjugation and PCR. RESULTS: The epidemiological studies showed that all six strains studied belonged to the same clone. Furthermore, all of them carried the same gene encoding for trimethoprim resistance (dfrA7), which was located in an integron within a chromosome. CONCLUSION: Continuous epidemiological surveillance is required to control the spread of pathogenic microorganisms and the dissemination of resistance-encoding genes among geographical areas.

Clonal dissemination of Yersinia enterocolitica strains with various susceptibilities to nalidixic acid.

Ten epidemiologically related Yersinia enterocolitica clinical isolates were studied. Six isolates were nalidixic acid resistant (MIC > 512 microg/ml), with mutations in the quinolone resistance-determining region (QRDR) of the gyrA gene, suggesting clonal dissemination of a nalidixic acid-susceptible Y. enterocolitica strain which has acquired different mutations generating resistance to nalidixic acid.

Distribution of beta-lactamases in Acinetobacter baumannii clinical isolates and the effect of Syn 2190 (AmpC inhibitor) on the MICs of different beta-lactam antibiotics.

The distribution of beta-lactamases in a group of 20 epidemiologically well defined Acinetobacter baumannii clinical isolates and the in vitro activity of Syn 2190, a novel beta-lactamase AmpC inhibitor, were determined. Twenty-five per cent of the strains carried and expressed a TEM-type beta-lactamase, whereas 35% had an OXA-type beta-lactamase. In nine out of 11 (82%) ceftazidime-resistant and four out of 13 (30.7%) cefepime-resistant strains, the MIC of these beta-lactam antibiotics decreased when determined in the presence of Syn 2190. Thus, our results suggest that in a high percentage of A. baumannii clinical isolates the increased production of AmpC, in combination or not with other resistance mechanisms, contributes to the resistance pattern in A. baumannii to beta-lactams.

Haematological and biochemical indices in young African children: in search of reference intervals

Introduction: The reference intervals of haematological and biochemical indices currently used in Africa are derived from data collected from populations living in industrialized countries. Few studies have been performed in Africa questioning the validity of these values when applied to local African populations. Objective: To provide reference intervals of haematological [haemoglobin (Hb), white blood cells (WBC), haematocrit (Htc) and platelets] and biochemical indices (ALT, creatinine and bilirubin) for children aged 1-4 from a rural area of southern Mozambique. Methods: Reference intervals were developed using the 2.5 and 97.5 centiles. Partition tests were performed to evaluate age and gender differences. Quantile regression models were estimated for those variables in which age partition was recommended. Deviances from linearity in the estimated models were evaluated using fractional polynomials of first or second degree. Agreement to classify normality, using the estimated reference intervals or values in use in a western paediatric hospital, was made using the kappa statistic. Results: Reference intervals for Hb, WBC, Htc, platelets, ALT and creatinine show significant differences by age. Gender differences were observed for creatinine values, while for bilirubin there were no significant differences for age or gender. Estimated Hb and Htc reference intervals in African children were lower than the accepted western ones, while ALT values were higher in the former. Agreement between normal classification, using the estimated intervals or the western values, was from slight to fai...

Relationship between haemoglobin and haematocrit in the definition of anaemia

Introduction: Anaemia is the most frequent haematological disorder in childhood. The notion that defines naemia does not change throughout life, although parameters used for its evaluation show significant variations during childhood. Haematocrit (Hct) (%) is usually defined as three times the value of haemoglobin (Hgb) (g/dl), while the clinical definition of anaemia is related to either an abnormal Hct or Hgb value. Objective: To evaluate the agreement between Hgb and Hct values in the definition of anaemia, the relationship between these two parameters and their age-dependence. Methods: The Hct and Hgb paired values from children aged 2-18 months from Ifakara (Tanzania) and children aged 1-4 years from Manhiça (Mozambique) were analysed. Haematological determinations of the Manhiça samples were done using a KX-21N cell counter (Kobe, Japan) and Ifakara samples were analysed in a semiautomatic cell counter (Sysmex F800 microcell counter, TOA Medical Electronics, Kobe, Japan). The kappa-statistic was used to calculate the agreement between anaemia definitions in each group. Crude and multivariate relationship between Hct and Hgb levels were analysed by linear regression model estimation. The age-dependence of the crude ratio (Hct/Hgb) was analysed using linear regression models and fractional polynomials. Results: The prevalences of mild and moderate anaemia as defined by Hgb levels in the Manhiça group were 61% and 6%, respectively, and 41% and 2% by Hct. In the Ifakara group these were 74% and 10%, respectively, by Hgb and 42% and 3% by Hct, respectively. Agreement between mild and moderate anaemia definitions made up from Hgb or from Hct levels were from fair to moderate. Hct levels decreased with age for high Hgb levels, whereas they increased for low Hgb levels. The classification of cases is improved when higher age-related cut-off values for Hct are used. The crude relationship between Hct and Hgb levels was significantly different from 3, and this was modified by age. The evaluation of the age-dependence ratio (Hct/Hgb) showed a non-linear relationship with an asymptotic trend to 3...

Analysis of the clonal relationship among clinical isolates of Salmonella enterica serovar Infantis by different typing methods

Salmonella Infantis has been the second most common serovar in Argentina in the last two years, being isolated mostly from paediatric hospitalised patients. In order to determine the clonal relationship among Salmonella Infantis strains, we examined 15 isolates from paediatric patient faeces in Argentina (12 geographically related and 3 geographically non-related) by using antimicrobial susceptibility, plasmid profiling, repetitive extragenic palindromic (REP) PCR, enterobacterial repetitive intergenic consensus (ERIC) PCR, and low-frequency restriction analysis of chromosomal DNA by pulsed field gel electrophoresis (PFGE). Four Spanish strains were included as controls of clonal diversity in molecular techniques. Antibiotype and plasmid profile was not useful as epidemiological tools. PFGE and REP-PCR were able to discriminate between Argentinean and Spanish isolates of Salmonella Infantis allowing to detect genetically related strains in three different cities. This finding indicates that a possible spread of a clone of this serovar in the North-eastern Region of Argentina has taken place in 1998

Efficacy of the RTS, S/AS02A vaccine against Plasmodium falciparum infection and disease in young African children: randomised controlled trial

Background: Development of an effective malaria vaccine could greatly contribute to disease control. RTS,S/AS02A is a pre-erythrocytic vaccine candidate based on Plasmodium falciparum circumsporozoite surface antigen. We aimed to assess vaccine efficacy, immunogenicity, and safety in young African children. Methods: We did a double-blind, phase IIb, randomised controlled trial in Mozambique in 2022 children aged 1-4 years. The study included two cohorts of children living in two separate areas which underwent different follow-up schemes. Participants were randomly allocated three doses of either RTS,S/AS02A candidate malaria vaccine or control vaccines. The primary endpoint, determined in cohort 1 (n=1605), was time to first clinical episode of P falciparum malaria (axillary temperature > or =37.5 degrees C and P falciparum asexual parasitaemia >2500 per microL) over a 6-month surveillance period. Efficacy for prevention of new infections was determined in cohort 2 (n=417). Analysis was per protocol. Findings: 115 children in cohort 1 and 50 in cohort 2 did not receive all three doses and were excluded from the per-protocol analysis. Vaccine efficacy for the first clinical episodes was 29.9% (95% CI 11.0-44.8; p=0.004). At the end of the 6-month observation period, prevalence of P falciparum infection was 37% lower in the RTS,S/AS02A group compared with the control group (11.9% vs 18.9%; p=0.0003). Vaccine efficacy for severe malaria was 57.7% (95% CI 16.2-80.6; p=0.019). In cohort 2, vaccine efficacy for extending time to first infection was 45.0% (31.4-55.9; p<0.0001). Interpretation: The RTS,S/AS02A vaccine was safe, well tolerated, and immunogenic. Our results show development of an effective vaccine against malaria is feasible