Prevalence and assemblage of Giardia duodenalis in a case-control study of children under 5 years from Jimma, Southwest Ethiopia.

Giardia duodenalis is a common pathogenic intestinal protozoan parasite with high prevalence in developing countries, especially among children. The distribution of giardia assemblages among humans and their clinical relevance remains controversial. This study aimed to determine the prevalence and assemblage of Giardia among children under 5 years of age in Jimma, Southwest Ethiopia. Employing a case-control design, 606 children presenting with diarrhea at Jimma university medical center and Serbo Health Center were enrolled from December 2016 to July 2018 along with 617 matched controls without diarrhea. Giardia was detected and typed using real-time PCR. Univariate and multivariate regression analysis was performed. The total prevalence of Giardia was 41% (501/1223) and did not differ significantly between cases and controls (40% vs 42%). Prevalence increased by age, with the highest prevalence seen in children aged ≥ 25 months. Children without diarrhea with a history of diarrhea during the last month were more likely to be Giardia positive compared to children with no history diarrhea (OR 1.8 and 95%CI; 1.1-2.9). Regardless of current diarrhea symptoms, assemblage B predominated with 89%, followed by assemblage A (8%) and mixed infection assemblage A and B (3%). We report a high prevalence of Giardia by PCR detection in Jimma, Ethiopia, with assemblage B being predominant. There was a similar distribution of Giardia assemblages between children with and without diarrhea. Increasing age was a risk factor for Giardia infection. Community-based prevention and control strategies need to be employed to decrease the risk of giardia infection.

Fluoroquinolone resistance among fecal extended spectrum ßeta lactamases positive Enterobacterales isolates from children in Dar es Salaam, Tanzania.

BACKGROUND: Fluoroquinolones have been, and continue to be, routinely used for treatment of many bacterial infections. In recent years, most parts of the world have reported an increasing trend of fluoroquinolone resistant (FQR) Gram-negative bacteria. METHODS: A cross-sectional study was conducted between March 2017 and July 2018 among children admitted due to fever to referral hospitals in Dar es Salaam, Tanzania. Rectal swabs were used to screen for carriage of extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-PE). ESBL-PE isolates were tested for quinolone resistance by disk diffusion method. Randomly selected fluroquinolone resistant isolates were characterized by using whole genome sequencing. RESULTS: A total of 142 ESBL-PE archived isolates were tested for fluoroquinolone resistance. Overall phenotypic resistance to ciprofloxacin, levofloxacin and moxifloxacin was found in 68% (97/142). The highest resistance rate was seen among Citrobacter spp. (100%, 5/5), followed by Klebsiella. pneumoniae (76.1%; 35/46), Escherichia coli (65.6%; 42/64) and Enterobacter spp. (31.9%; 15/47). Whole genome sequencing (WGS) was performed on 42 fluoroquinolone resistant-ESBL producing isolates and revealed that 38/42; or 90.5%, of the isolates carried one or more plasmid mediated quinolone resistance (PMQR) genes. The most frequent PMQR genes were aac(6')-lb-cr (74%; 31/42), followed by qnrB1 (40%; 17/42), oqx, qnrB6 and qnS1. Chromosomal mutations in gyrA, parC and parE were detected among 19/42 isolates, and all were in E. coli. Most of the E. coli isolates (17/20) had high MIC values of > 32 µg/ml for fluoroquinolones. In these strains, multiple chromosomal mutations were detected, and all except three strains had additional PMQR genes. Sequence types, ST131 and ST617 predominated among E. coli isolates, while ST607 was more common out of 12 sequence types detected among the K. pneumoniae. Fluoroquinolone resistance genes were mostly associated with the IncF plasmids. CONCLUSION: The ESBL-PE isolates showed high rates of phenotypic resistance towards fluoroquinolones likely mediated by both chromosomal mutations and PMQR genes. Chromosomal mutations with or without the presence of PMQR were associated with high MIC values in these bacteria strains. We also found a diversity of PMQR genes, sequence types, virulence genes, and plasmid located antimicrobial resistance (AMR) genes towards other antimicrobial agents.

Genetic determinants of macrolide and tetracycline resistance in penicillin non-susceptible Streptococcus pneumoniae isolates from people living with HIV in Dar es Salaam, Tanzania.

BACKGROUND: Over one million yearly deaths are attributable to Streptococcus pneumoniae and people living with HIV are particularly vulnerable. Emerging penicillin non-susceptible Streptococcus pneumoniae (PNSP) challenges therapy of pneumococcal disease. The aim of this study was to determine the mechanisms of antibiotic resistance among PNSP isolates by next generation sequencing. METHODS: We assessed 26 PNSP isolates obtained from the nasopharynx from 537 healthy human immunodeficiency virus (HIV) infected adults in Dar es Salaam, Tanzania, participating in the randomized clinical trial CoTrimResist (ClinicalTrials.gov identifier: NCT03087890, registered on 23rd March, 2017). Next generation whole genome sequencing on the Illumina platform was used to identify mechanisms of resistance to antibiotics among PNSP. RESULTS: Fifty percent (13/26) of PNSP were resistant to erythromycin, of these 54% (7/13) and 46% (6/13) had MLSB phenotype and M phenotype respectively. All erythromycin resistant PNSP carried macrolide resistance genes; six isolates had mef(A)-msr(D), five isolates had both erm(B) and mef(A)-msr(D) while two isolates carried erm(B) alone. Isolates harboring the erm(B) gene had increased MIC (> 256 µg/mL) towards macrolides, compared to isolates without erm(B) gene (MIC 4-12 µg/mL) p < 0.001. Using the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines, the prevalence of azithromycin resistance was overestimated compared to genetic correlates. Tetracycline resistance was detected in 13/26 (50%) of PNSP and all the 13 isolates harbored the tet(M) gene. All isolates carrying the tet(M) gene and 11/13 isolates with macrolide resistance genes were associated with the mobile genetic element Tn6009 transposon family. Of 26 PNSP isolates, serotype 3 was the most common (6/26), and sequence type ST271 accounted for 15% (4/26). Serotypes 3 and 19 displayed high-level macrolide resistance and frequently carried both macrolide and tetracycline resistance genes. CONCLUSION: The erm(B) and mef(A)-msr(D) were common genes conferring resistance to MLSB in PNSP. Resistance to tetracycline was conferred by the tet(M) gene. Resistance genes were associated with the Tn6009 transposon.

Gastrointestinal colonization of extended-spectrum beta-lactamase-producing bacteria among children below five years of age hospitalized with fever in Dar es Salaam, Tanzania.

OBJECTIVES: Gastrointestinal colonization of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-PE) is of concern because prior colonization increases risk for subsequent infections. To date, the link between ESBL-PE faecal carriage and the risk of subsequent ESBL-PE infection has not been well established, and information on carriage of such pathogens among children with invasive infections such as bloodstream infections (BSI) remains to be explored worldwide. METHODS: This cross-sectional study was conducted among children under the age of 5 years admitted for febrile illness in Dar es Salaam, Tanzania, between March 2017 and July 2018. We used rectal swabs to screen for ESBL-PE using selective media, ChromID ESBL. Bacterial isolates were identified by MALDI-TOF. Blood cultures were drawn from all children. Antimicrobial susceptibility testing was done using a disk diffusion method. ESBL alleles were identified by real-time PCR and sequencing. RESULTS: The overall prevalence of ESBL-PE carriage was 56% (112/200) and was highest among children 4 to 6 months old (17/21, 81%) (P = 0.05). Children with BSI had high ESBL-PE carriage (78.4%) compared to those without BSI (53.1%) (P = 0.02; aOR 3.4, 95% confidence interval 1.20-9.58). The most common isolate was E. coli (64/112, 45%). Sixteen pairs of ESBL-PE isolates (from the gut and from blood) had a similar antimicrobial susceptibility profile. We detected blaCTX-M gene in 97% of all phenotypically detected ESBL-PE; among those, blaCTX-M-15 was dominant (99%). CONCLUSION: We report a high prevalence of ESBL-PE faecal carriage among children with BSI in Tanzania. Colonization of ESBL-PE was a risk factor for ESBL-BSI.

First identification of bla NDM-5 producing Escherichia coli from neonates and a HIV infected adult in Tanzania.

Introduction. Carbapenem-resistant members of the family Enterobacteriaceae are emerging as a global public-health threat and cause substantial challenges in clinical practice.Gap Statement. There is a need for increased and continued genomic surveillance of antimicrobial resistance genes globally in order to detect outbreaks and dissemination of clinically important resistance genes and their associated mobile genetic elements in human pathogens.Aim. To describe the resistance mechanisms of carbapenem-resistant Escherichia coli.Methods. Rectal swabs from neonates and newly diagnosed human immunodeficiency virus (HIV) infected adults were collected between April 2017 and May 2018 and screened for faecal carriage of carbapenamases and OXA-48 producing members of the family Enterobacteriaceae. Bacterial isolates were identified using matrix assisted laser desorption ionization time of flight mass spectrometry. Antimicrobial susceptibility testing was performed by E-test. Whole genomes of carbapenem-resistant E. coli were investigated using a hybrid assembly of Illumina and Oxford Nanopore Technologies sequencing reads.Results. Three carbapenem-resistant E. coli were detected, two from neonates and one from an HIV infected adult. All three isolates carried bla NDM-5. Two E. coli from neonates belonged to ST167 and bla NDM-5 co-existed with bla CTX-M-15 and bla OXA-01, and all were carried on IncFIA type plasmids. The E. coli from the HIV infected adult belonged to ST2083, and carried bla NDM-5 on an IncX3 type plasmid and bla CMY-42 on an IncI type plasmid. All bla NDM-5 carrying plasmids contained conjugation related genes. In addition, E. coli from the HIV infected adult carried three more plasmid types; IncFIA, IncFIB and Col(BS512). One E. coli from a neonate also carried one extra plasmid Col(BS512). All three E. coli harboured resistance genes to fluoroquinolone, aminoglycosides, sulfamethoxazole, trimethoprim, macrolides and tetracycline, carried on the IncFIA type plasmid. Furthermore, E. coli from the neonates carried a chloramphenicol resistance gene (catB3), also on the IncFIA plasmid. All three isolates were susceptible to colistin.Conclusion. This is the first report, to our knowledge, from Tanzania detecting bla NDM-5 producing E. coli. The carbapenemase gene was carried on an IncFIA and IncX3 type plasmids. Our findings highlight the urgent need for a robust antimicrobial resistance (AMR) surveillance system to monitor and rapidly report on the incidence and spread of emerging resistant bacteria in Tanzania.

Predominance of PVL-negative community-associated methicillin-resistant Staphylococcus aureus sequence type 8 in newly diagnosed HIV-infected adults, Tanzania.

Difficult-to-treat infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are of concern in people living with HIV infection as they are more vulnerable to infection. We aimed to identify molecular characteristics of MRSA colonizing newly diagnosed HIV-infected adults in Tanzania. Individuals newly diagnosed with HIV infection were recruited in Dar es Salaam, Tanzania, from April 2017 to May 2018, as part of the randomized clinical trial CoTrimResist ( ClinicalTrials.gov identifier: NCT03087890). Nasal/nasopharyngeal isolates of Staphylococcus aureus were susceptibility tested by disk diffusion method, and cefoxitin-resistant isolates were characterized by short-reads whole genome sequencing. Four percent (22/537) of patients carried MRSA in the nose/nasopharynx. MRSA isolates were frequently resistant towards gentamicin (95%), ciprofloxacin (91%), and erythromycin (82%) but less often towards trimethoprim-sulfamethoxazole (9%). Seventy-three percent had inducible clindamycin resistance. Erythromycin-resistant isolates harbored ermC (15/18) and LmrS (3/18) resistance genes. Ciprofloxacin resistance was mediated by mutations of the quinolone resistance-determining region (QRDR) sequence in the gyrA (S84L) and parC (S80Y) genes. All isolates belonged to the CC8 and ST8-SCCmecIV MRSA clone. Ninety-five percent of the MRSA isolates were spa-type t1476, and one exhibited spa-type t064. All isolates were negative for Panton-Valentine leucocidin (PVL) and arginine catabolic mobile element (ACME) type 1. All ST8-SCCmecIV-spa-t1476 MRSA clones from Tanzania were unrelated to the globally successful USA300 clone. Carriage of ST8 MRSA (non-USA300) was common among newly diagnosed HIV-infected adults in Tanzania. Frequent co-resistance to non-beta lactam antibiotics limits therapeutic options when infection occurs.

Molecular characterisation of the first New Delhi metallo-ß-lactamase 1-producing Acinetobacter baumannii from Tanzania.

BACKGROUND: We aimed to characterise the genetic determinants and context of two meropenem-resistant clinical isolates of Acinetobacter baumannii isolated from children hospitalised with bloodstream infections in Dar es Salaam, Tanzania. METHODS: Antimicrobial susceptibility was determined by disc diffusion E-test and broth microdilution. Genomes were completed using a hybrid assembly of Illumina and Oxford Nanopore Technologies sequencing reads and characterisation of the genetic context of resistance genes, multi-locus sequence types (STs) and phylogenetic analysis was determined bioinformatically. RESULTS: Twelve A. baumannii were isolated from 2226 blood cultures, two of which were meropenem-resistant. The two meropenem-resistant isolates, belonging to distinct STs, ST374 and ST239, were found to harbour blaNDM-1, which was chromosomally located in isolate DT0544 and plasmid-located in isolate DT01139. The genetic environment of blaNDM-1 shows the association of insertion sequence ISAba125 with blaNDM-1 in both isolates. Both isolates also harboured genes conferring resistance to other ß-lactams, aminoglycosides and cotrimoxazole. CONCLUSIONS: This is the first report of New Delhi metallo-ß-lactamase-producing isolates of A. baumannii from Tanzania. The genetic context of blaNDM-1 provides further evidence of the importance of ISAba125 in the spread of blaNDM-1 in A. baumannii. Local surveillance should be strengthened to keep clinicians updated on the incidence of these and other multidrug-resistant and difficult-to-treat bacteria.

Bacteraemia, Malaria, and Case Fatality Among Children Hospitalized With Fever in Dar es Salaam, Tanzania.

BACKGROUND: Febrile illness is the commonest cause of hospitalization in children <5 years in sub-Saharan Africa, and bacterial bloodstream infections and malaria are major causes of death. METHODS: From March 2017 to July 2018, we enrolled 2,226 children aged 0-5 years hospitalized due to fever in four major public hospitals of Dar es Salaam, namely, Amana, Temeke, and Mwananyamala Regional Hospitals and Muhimbili National Hospital. We recorded social demographic and clinical data, and we performed blood-culture and HIV-antibody testing. We used qPCR to quantify Plasmodium falciparum parasitaemia and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) to identify bacterial isolates. Disk diffusion method was used for antimicrobial susceptibility testing. RESULTS: Nineteen percent of the children (426/2,226) had pathogens detected from blood. Eleven percent (236/2,226) of the children had bacteraemia/fungaemia and 10% (204/2,063) had P. falciparum malaria. Ten children had concomitant malaria and bacteraemia. Gram-negative bacteria (64%) were more frequent than Gram-positive (32%) and fungi (4%). Over 50% of Gram-negative bacteria were extended-spectrum beta-lactamase (ESBL) producers and multidrug resistant. Methicillin resistant Staphylococcus aureus (MRSA) was found in 11/42 (26.2%). The most severe form of clinical malaria was associated with high parasitaemia (>four million genomes/µL) of P. falciparum in plasma. Overall, in-hospital death was 4% (89/2,146), and it was higher in children with bacteraemia (8%, 18/227) than malaria (2%, 4/194, p = 0.007). Risk factors for death were bacteraemia (p = 0.03), unconsciousness at admission (p < 0.001), and admission at a tertiary hospital (p = 0.003). CONCLUSION: Compared to previous studies in this region, our study showed a reduction in malaria prevalence, a decrease in in-hospital mortality, and an increase in antimicrobial resistance (AMR) including ESBLs and multidrug resistance. An increase of AMR highlights the importance of continued strengthening of diagnostic capability and antimicrobial stewardship programs. We also found malaria and bacteraemia contributed equally in causing febrile illness, but bacteraemia caused higher in-hospital death. The most severe form of clinical malaria was associated with P. falciparum parasitaemia.

Plasma parasitemia as assessed by quantitative PCR in relation to clinical disease severity in African adults with falciparum malaria with and without HIV co-infection.

PURPOSE: When considering malaria disease severity, estimation of parasitemia in erythrocytes is important, but sometimes misleading, since the infected erythrocytes may be sequestered in peripheral capillaries. In African children and Asian adults with falciparum malaria, parasitemia as assessed by quantitative PCR (qPCR) in plasma seems to be a valuable indicator of disease severity, but data on African adults as well as the impact of co-infection with HIV is lacking. METHODS: In 131 patients with falciparum malaria in a public tertiary teaching hospital in Mozambique, plasma malaria parasitemia as assessed by qPCR, compared to qualitative malaria PCR in blood cell fraction, was related to malaria disease severity and HIV co-infection. RESULTS: Of the 131 patients with falciparum malaria, based on positive qualitative PCR in the blood cell fraction, 93 patients (72%) had positive malaria qPCR in plasma. Patients with severe malaria as defined by the WHO criteria had higher malaria quantitative plasma parasitemia (median 143 genomes/µL) compared to those with uncomplicated malaria (median 55 genomes/µL, p = 0.037) in univariate analysis, but this difference was attenuated after adjusting for age, sex and HIV co-infection (p = 0.055). A quarter of the patients with severe malaria had negative qPCR in plasma. CONCLUSIONS: This study of adult African in-patients with falciparum malaria with and without HIV co-infection, neither confirms nor rejects previous studies of malaria qPCR in plasma as an indicator of disease severity in patients with falciparum malaria. There is a need for further and larger studies to clarify if parasitemia as assessed malaria qPCR in plasma could be a surrogate marker of disease severity in falciparum malaria.

Nasal Carriage of Methicillin-Resistant Staphylococcus aureus among Health Care Workers in Tertiary and Regional Hospitals in Dar es Salam, Tanzania.

Methicillin-resistant Staphylococcus aureus (MRSA) among health care workers (HCWs) increases the risk of spreading the organism in hospital settings. A cross-sectional study was conducted between June and October 2016 among HCWs in tertiary and regional hospitals in Dar es Salaam, Tanzania, to determine the MRSA nasal carriage rate. Nasal swabs were collected from HCWs and cultured on mannitol salt agar. S. aureus was identified based on colonial morphology, Gram staining, catalase, coagulase, and DNase test results. MRSA was detected using the cefoxitin disk. Among 379 HCWs enrolled, 157/379 (41.4%) were colonized with S. aureus, of whom 59 (37.6%) were MRSA carriers giving an overall prevalence of 59/379 (15.6%). MRSA carriage was high among HCWs in Temeke (56.9%) and Amana (37.5%) regional hospitals. A high proportion of MRSA carriage was detected among nurses (35, 45.5%). MRSA isolates showed high resistance toward kanamycin (83.7%), gentamicin (83.1%), ciprofloxacin (71.2%), and trimethoprim-sulphamethoxazole (46.8%) compared to methicillin-sensitive S. aureus isolates (p ≤ 0.001). In conclusion, we found a high nasal carriage of MRSA and resistance to commonly prescribed antimicrobial agents among HCWs. Implementation of infection control measures including contact precautions, urgent reporting of MRSA laboratory results, and routine MRSA screening of HCWs is highly needed to reduce MRSA spreading.

Erratum to: Prevalence of methicillin-resistant Staphylococcus aureus carriage on admission among patients attending regional hospitals in Dar es Salaam, Tanzania.

Following publication of the original article [1], author Elia Mmbaga pointed out that her name had been misspelt as Elia Mbaga.

Prevalence of methicillin-resistant Staphylococcus aureus carriage on admission among patients attending regional hospitals in Dar es Salaam, Tanzania.

BACKGROUND: Methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogen responsible for hospital and community acquired infection. Colonization with MRSA is associated with a high risk of developing infection. This study aimed to determine the rate of MRSA carriage on admission and the associated risk factors among patients attending regional hospitals, in Dar es Salaam, Tanzania. RESULTS: A total of 258 patients were included in this study. Nasal swabs were collected on admission to the hospital and after 48 h of hospital stay for detection of MRSA. Of 258 patients enrolled, 89 (34.5%) were colonized with S. aureus and out them 22 (24.7%) were carriers of MRSA, giving an overall MRSA nasal carriage rate of 8.5% (22/258). One patient acquired MRSA while admitted in the hospital. Most of the S. aureus isolates 85 (95.5%) were resistant to penicillin. Resistance to gentamycin, ciprofloxacin, kanamycin, linezolid and mupirocin were 14.6, 11.2, 11.2, 3.4 and 1.1%, respectively. The prevalence of inducible clindamycin resistance, constitutive clindamycin resistance, MS phenotype (resistance to erythromycin alone), and multidrug resistance was 21.3, 3.4, 12.4, and 16.9%, respectively. We observed a statistically significant association between MRSA and multiple drugs resistance among S. aureus isolates (p = 0.001). Of the risk factors investigated none were statistically significant associated with MRSA. CONCLUSION: There is a high prevalence of MRSA among patients on admission at the two municipal hospitals in Dar es Salaam. The high prevalence of MRSA and the increased rates of resistance to commonly used antimicrobials among MRSA isolates call for attention to the importance of including the screening of MRSA in our hospitals setting in order to prevent further spread of MRSA strains to other patients and to the communities. Control and prevention strategies should be emphasized including decolonization.

Comprehensive Analysis of Prevalence, Epidemiologic Characteristics, and Clinical Characteristics of Monoinfection and Coinfection in Diarrheal Diseases in Children in Tanzania.

The role of interactions between intestinal pathogens in diarrheal disease is uncertain. From August 2010 to July 2011, we collected stool samples from 723 children admitted with diarrhea (cases) to 3 major hospitals in Dar es Salaam, Tanzania, and from 564 nondiarrheic children (controls). We analyzed the samples for 17 pathogens and assessed interactions between coinfections in additive and multiplicative models. At least one pathogen was detected in 86.9% of the cases and 62.8%, of the controls. Prevalence of coinfections was 58.1% in cases and 40.4% in controls. Rotavirus, norovirus genogroup II, Cryptosporidium, and Shigella species/enteroinvasive Escherichia coli were significantly associated with diarrhea both as monoinfections and as coinfections. In the multiplicative interaction model, we found 2 significant positive interactions: rotavirus + Giardia (odds ratio (OR) = 23.91, 95% confidence interval (CI): 1.21, 470.14) and norovirus GII + enteroaggregative E. coli (OR = 3.06, 95% CI: 1.17, 7.98). One significant negative interaction was found between norovirus GII + typical enteropathogenic E. coli (OR = 0.09, 95% CI: 0.01, 0.95). In multivariate analysis, risk factors for death were presence of blood in stool and severe dehydration. In conclusion, coinfections are frequent, and the pathogenicity of each organism appears to be enhanced by some coinfections and weakened by others. Severity of diarrhea was not affected by coinfections.

Detection of CTX-M-15 beta-lactamases in Enterobacteriaceae causing hospital- and community-acquired urinary tract infections as early as 2004, in Dar es Salaam, Tanzania.

BACKGROUND: The spread of Extended Spectrum ß-lactamases (ESBLs) among Enterobacteriaceae and other Gram-Negative pathogens in the community and hospitals represents a major challenge to combat infections. We conducted a study to assess the prevalence and genetic makeup of ESBL-type resistance in bacterial isolates causing community- and hospital-acquired urinary tract infections. METHODS: A total of 172 isolates of Enterobacteriaceae were collected in Dar es Salaam, Tanzania, from patients who met criteria of community and hospital-acquired urinary tract infections. We used E-test ESBL strips to test for ESBL-phenotype and PCR and sequencing for detection of ESBL genes. RESULTS: Overall 23.8% (41/172) of all isolates were ESBL-producers. ESBL-producers were more frequently isolated from hospital-acquired infections (32%, 27/84 than from community-acquired infections (16%, 14/88, p < 0.05). ESBL-producers showed high rate of resistance to ciprofloxacin (85.5%), doxycycline (90.2%), gentamicin (80.5%), nalidixic acid (84.5%), and trimethoprim-sulfamethoxazole (85.4%). Furthermore, 95% of ESBL-producers were multi-drug resistant compared to 69% of non-ESBL-producers (p < 0.05). The distribution of ESBL genes were as follows: 29/32 (90.6%) bla CTX-M-15, two bla SHV-12, and one had both bla CTX-M-15 and bla SHV-12. Of 29 isolates carrying bla CTX-M-15, 69% (20/29) and 31% (9/29) were hospital and community, respectively. Bla SHV-12 genotypes were only detected in hospital-acquired infections. CONCLUSION: bla CTX-M-15 is a predominant gene conferring ESBL-production in Enterobacteriaceae causing both hospital- and community-acquired infections in Tanzania.

Molecular Characterization of Cotrimoxazole Resistance Genes and Their Associated Integrons in Clinical Isolates of Gram-Negative Bacteria from Tanzania.

Cotrimoxazole is widely used, particularly as a prophylactic drug in HIV patients. We assessed resistance mechanisms among cotrimoxazole resistant-Gram negative bacterial isolates (n = 123) obtained from blood (n = 69) and urine (n = 54) from Tanzanian patients. sul genes were detected in 98% (121/123) of the isolates. Coexistence of sul1 and sul2 was common (49/123). The dfr genes were found in 63% (77/123) of all isolates. sul1, dfrA15, and dfrA5 genes predominated among Klebsiella pneumoniae, while sul2 and dfrA1 genes were frequent in Escherichia coli isolates. Two isolates, both K. pneumoniae, carried sul3. Integrons were detected in 81.3% (100/123) of all isolates. Class 1 integrons were found in 95% (42/44), 53% (23/43), and 80.6% (25/31) of K. pneumoniae, E. coli, and other Enterobacteriaceae isolates, respectively. Class 2 integrons were found in 14% of E. coli, but not in K. pneumoniae. All sul1 genes in K. pneumoniae were carried in class 1 integrons. Gene cassette arrays dfrA5 and dfrA15-aadA1 were most frequently associated with class 1 integrons, while class 2 integrons contained only dfrA1-sat2-aadA1 gene cassettes. This is the first report of sul3 gene in K. pneumoniae from human sources. The finding that mechanisms differ between E. coli and K. pneumoniae may broaden our understanding of cotrimoxazole resistance.

High Prevalence of Faecal Carriage of ESBL-Producing Enterobacteriaceae among Children in Dar es Salaam, Tanzania.

BACKGROUND: Faecal carriage of ESBL-producing bacteria is a potential risk for transmission and infection. Little is known about faecal carriage of antibiotic resistance in Tanzania. This study aimed to investigate the prevalence of faecal carriage of ESBL-producing Enterobacteriaceae and to identify risk factors for carriage among young children in Tanzania. METHODOLOGY/PRINCIPAL FINDINGS: From August 2010 to July 2011, children below 2 years of age were recruited in Dar es Salaam, including healthy community children (n = 250) and children hospitalized due to diarrhoea (n = 250) or other diseases (n = 103). ChromID ESBL agar and ChromID CARBA SMART agar were used for screening. Antimicrobial susceptibility testing was performed by the disk diffusion method. ESBL genotypes were identified by Real-Time PCR and sequencing. The overall prevalence of ESBL carriage was 34.3% (207/ 603). The prevalence of ESBL carriage was significantly higher among hospitalized children (50.4%), compared to community children (11.6%; P < 0.001; OR = 7.75; 95% CI: 4.99-12.03). We found high prevalence of Multidrug-resistance (94%) among Escherichia coli and Klebsiella pneumoniae isolates. No resistance to carbapenems was detected. For the majority of isolates (94.7%) we detected a blaCTX-M-15-like gene. In addition, the plasmid mediated AmpC beta-lactamase CMY-2 was detected for the first time in Tanzania. ESBL prevalence was significantly higher among HIV positive (89.7%) than HIV negative (16.9%) children (P = 0.001; OR = 9.99; 95% CI: 2.52-39.57). Use of antibiotics during the past 14 days and age below 1 year was also associated with ESBL carriage. CONCLUSIONS/SIGNIFICANCE: We report a high rate of faecal carriage of ESBL-producing Enterobacteriaceae among children below 2 years of age in Tanzania, particularly those with HIV-infection. Resistance to a majority of the available antimicrobials commonly used for children in Tanzania leaves few treatment options for infections when caused by these bacteria.

Prevalence of Cryptosporidium parvum/hominis, Entamoeba histolytica and Giardia lamblia among Young Children with and without Diarrhea in Dar es Salaam, Tanzania.

BACKGROUND: Although enteroparasites are common causes of diarrheal illness, few studies have been performed among children in Tanzania. This study aimed to investigate the prevalence of Cryptosporidium parvum/hominis, Entamoeba histolytica and Giardia lamblia among young children in Dar es Salaam, Tanzania, and identify risk factors for infection. METHODOLOGY/PRINCIPAL FINDINGS: We performed an unmatched case-control study among children < 2 years of age in Dar es Salaam, recruited from August 2010 to July 2011. Detection and identification of protozoans were done by PCR techniques on DNA from stool specimens from 701 cases of children admitted due to diarrhea at the three study hospitals, and 558 controls of children with no history of diarrhea during the last month prior to enrollment. The prevalence of C. parvum/hominis was 10.4% (84.7% C. hominis), and that of G. lamblia 4.6%. E. histolytica was not detected. The prevalence of Cryptosporidium was significantly higher in cases (16.3%) than in controls (3.1%; P < 0.001; OR = 6.2; 95% CI: 3.7-10.4). G. lamblia was significantly more prevalent in controls (6.1%) than in cases (3.4%; P = 0.027; OR = 1.8; 95% CI: 1.1-3.1). Cryptosporidium infection was found more often in HIV-positive (24.2%) than in HIV-negative children (3.9%; P < 0.001; OR = 7.9; 95% CI: 3.1-20.5), and was also associated with rainfall (P < 0.001; OR = 2.41; 95% CI: 1.5-3.8). Among cases, stunted children had significantly higher risk of being infected with Cryptosporidium (P = 0.011; OR = 2.12; 95% CI: 1.2-3.8). G. lamblia infection was more prevalent in the cool season (P = 0.004; OR = 2.2; 95% CI: 1.3-3.8), and more frequent among cases aged > 12 months (P = 0.003; OR = 3.5; 95% CI: 1.5-7.8). Among children aged 7-12 months, those who were breastfed had lower prevalence of G. lamblia infection than those who had been weaned (P = 0.012). CONCLUSIONS: Cryptosporidium infection is common among young Tanzanian children with diarrhea, particularly those living with HIV, and infection is more frequent during the rainy season. G. lamblia is frequently implicated in asymptomatic infections, but rarely causes overt diarrheal illness, and its prevalence increases with age.

Genetic diversity of circulating rotavirus strains in Tanzania prior to the introduction of vaccination.

BACKGROUND: Tanzania currently rolls out vaccination against rotavirus-diarrhea, a major cause of child illness and death. As the vaccine covers a limited number of rotavirus variants, this study describes the molecular epidemiology of rotavirus among children under two years in Dar es Salaam, Tanzania, prior to implementation of vaccination. METHODS: Stool specimens, demographic and clinical information, were collected from 690 children admitted to hospital due to diarrhea (cases) and 545 children without diarrhea (controls) during one year. Controls were inpatient or children attending child health clinics. Rotavirus antigen was detected using ELISA and positive samples were typed by multiplex semi-nested PCR and sequencing. RESULTS: The prevalence of rotavirus was higher in cases (32.5%) than in controls (7.7%, P<0.001). The most common G genotypes were G1 followed by G8, G12, and G4 in cases and G1, G12 and G8 in controls. The Tanzanian G1 variants displayed 94% similarity with the Rotarix vaccine G1 variant. The commonest P genotypes were P[8], P[4] and P[6], and the commonest G/P combination G1 P[8] (n = 123), G8 P[4] and G12 P[6]. Overall, rotavirus prevalence was higher in cool (23.9%) than hot months (17.1%) of the year (P = 0.012). We also observed significant seasonal variation of G genotypes. Rotavirus was most frequently found in the age group of four to six months. The prevalence of rotavirus in cases was lower in stunted children (28.9%) than in non-stunted children (40.1%, P = 0.003) and lower in HIV-infected (15.4%, 4/26) than in HIV-uninfected children (55.3%, 42/76, P<0.001). CONCLUSION: This pre-vaccination study shows predominance of genotype G1 in Tanzania, which is phylogenetically distantly related to the vaccine strains. We confirm the emergence of genotype G8 and G12. Rotavirus infection and circulating genotypes showed seasonal variation. This study also suggests that rotavirus may not be an opportunistic pathogen in children infected with HIV.

Penicillin resistance and serotype distribution of Streptococcus pneumoniae in nasopharyngeal carrier children under 5 years of age in Dar es Salaam, Tanzania.

This study aimed to determine the magnitude of nasopharyngeal carriage, antimicrobial resistance and serotype distribution of Streptococcus pneumoniae in healthy children under 5 years of age in Tanzania. Nasopharyngeal swabs were obtained from 300 healthy children attending a child health clinic at Muhimbili National Hospital in Dar es Salaam, Tanzania. S. pneumoniae was isolated and identified using conventional methods. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disc diffusion method. Penicillin MICs and serotypes were determined by an agar gradient diffusion method and the Quellung reaction, respectively. A total of 105 samples (35 .0%) were positive for S. pneumoniae and 115 serotypes were detected (ten specimens yielded two serotypes each). Overall, 78 of 115 isolates (67.8 %) were penicillin-non-susceptible pneumococci (PNSP). The resistance levels of S. pneumoniae to trimethoprim-sulfamethoxazole, tetracycline, erythromycin, chloramphenicol and ceftriaxone were 82.6, 10.4, 6.0, 3.5 and 0.0 %, respectively. Multidrug resistance was detected in 19 isolates (16.5 %). The most prevalent serotypes were 19F (n = 25, 21.7 %), 6B (n = 15, 13.0 %), 9V (n = 14, 12.2 %) and 13 (n = 14, 12.2 %). Of the 64 pneumococcal isolates potentially covered by the seven-valent pneumococcal conjugate vaccine (PCV7), 44 (68.8 %) were PNSP. A high prevalence of PNSP, common pneumococcal serotypes circulating worldwide, was found, and many of the resistant pneumococci strains are covered by the PCV7. These findings indicate that the carriage rate of such resistant strains could be influenced by an appropriate vaccination programme in the study setting and by reinforcing regulations on the rational use of antimicrobial agents.

Age specific aetiological agents of diarrhoea in hospitalized children aged less than five years in Dar es Salaam, Tanzania.

BACKGROUND: This study aimed to determine the age-specific aetiologic agents of diarrhoea in children aged less than five years. The study also assessed the efficacy of the empiric treatment of childhood diarrhoea using Integrated Management of Childhood Illness (IMCI) guidelines. METHODS: This study included 280 children aged less than 5 years, admitted with diarrhoea to any of the four major hospitals in Dar es Salaam. Bacterial pathogens were identified using conventional methods. Enzyme Linked Immunosorbent Assay (ELISA) and agglutination assay were used to detect viruses and intestinal protozoa, respectively. Antimicrobial susceptibility was determined using Kirby-Bauer disk diffusion method. RESULTS: At least one of the searched pathogens was detected in 67.1% of the cases, and mixed infections were detected in 20.7% of cases. Overall, bacteria and viruses contributed equally accounting for 33.2% and 32.2% of all the cases, respectively, while parasites were detected in 19.2% patients. Diarrhoeagenic Escherichia coli (DEC) was the most common enteric pathogen, isolated in 22.9% of patients, followed by Cryptosporidium parvum (18.9%), rotavirus (18.1%) and norovirus (13.7%). The main cause of diarrhoea in children aged 0 to 6 months were bacteria, predominantly DEC, while viruses predominated in the 7-12 months age group. Vibrio cholerae was isolated mostly in children above two years. Shigella spp, V. cholerae and DEC showed moderate to high rates of resistance to erythromycin, ampicillin, chloramphenicol and tetracycline (56.2-100%). V. cholerae showed full susceptibility to co-trimoxazole (100%), while DEC and Shigella showed high rate of resistance to co-trimoxazole; 90.6% and 93.3% respectively. None of the bacterial pathogens isolated showed resistance to ciprofloxacin which is not recommended for use in children. Cefotaxime resistance was found only in 4.7% of the DEC. CONCLUSION: During the dry season, acute watery diarrhoea is the most common type of diarrhoea in children under five years in Dar es Salaam and is predominantly due to DEC, C. parvum, rotaviruses and noroviruses. Constant antibiotic surveillance is warranted as bacteria were highly resistant to various antimicrobial agents including co-trimoxazole and erythromycin which are currently recommended for empiric treatment of diarrhoea.