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This review focuses on the reports of salmonellosis by investigators in different parts of Ethiopia, in particular focusing on the levels of typhoid fever. Many of the reports are published in local journals that are not available online. There have been seven studies which diagnosed typhoid fever by laboratory culture and there is no coordinated epidemiological surveillance. All conducted research and reports from different health institutions in Ethiopia indicate that typhoid fever was still a common problem up to the most recent study in 2000 and that the extensive use of first-line drugs has led to the development of multiple drug resistance. In the sites covered by this review, the total number of published cases of typhoid fever dropped over time reflecting the decline in research capacity in the country. Data on the proportion of patients infected by different serovars of Salmonella suggest that the non-Typhi serovars of Salmonella are increasing. The published evidence suggests that typhoid fever is a current public health problem in Ethiopia although population based surveys, based on good microbiological diagnosis, are urgently needed. Only then can the true burden of enteric fever be estimated and the benefit of public health control measures, such as health education, safe water provision, improved food hygienic practices and eventually vaccination, be properly assessed.

Background: Expanded access to HIV therapy in the developing world raises serious concerns regarding the potential emergence and transmission of drug-resistant HIV strains. Although HIV drug resistance surveillance is recommended to track transmitted HIV drug resistance among newly infected individuals, the financial constraints in resource-limited countries prohibit such surveillance on a regular basis. The World Health Organization (WHO) recently introduced guidelines to address this issue., Methods: A survey was conducted in Ethiopia following the WHO guidelines to assess transmitted HIV drug resistance among recently HIV-infected individuals in Addis Ababa. Antiretroviral drug usage started 3 years earlier than commencement of the current expanded access to antiretroviral therapy in Ethiopia., Results: Of 75 eligible samples, 39 (52%) were successfully sequenced and genotyped in the protease and reverse transcriptase region, using both the ViroSeq and TrueGene genotyping systems, and analysed for drug resistance mutations using an algorithm from the Stanford HIV Reverse Transcriptase and Protease Database. The analysis revealed that transmitted HIV drug resistance in Addis Ababa is below the 5% threshold level for all three classes of antiretrovirals., Conclusions: The current first-line antiretroviral therapy strategy can be used with confidence in Ethiopia at this time; however, Ethiopia should conduct similar periodic surveys that include the capitals of Ethiopia's larger regional states to ensure early detection of any changes in the country's HIV drug resistance trend.

Objective: Evaluation and monitoring of Human Immunodeficiency Virus (HIV) testing reagents at the point of service is helpful to prevent the occurrence of problems related to testing and interpretation. To evaluate the implementation of HIV rapid test kits at the point of services in voluntarily counseling and testing (VCT) and diagnostic centers in Ethiopia., Methods: The assessment was the third phase of evaluation of HIV rapid test kits in Ethiopia followed from phase-I and phase-II. Known proficiency testing panels, well-structured questionnaire (addressing type of tests, human resource and problems related to tests), onsite supervision and retesting of samples collected from sites were used to evaluate the performances of reagents and laboratories., Results: Forty-four health institutions were included. Thirty-six (90.0%) health institutions had trained human resource on HIV testing. In 27 (61.4%) three types of HIV rapid test kits (Determine, Capillus and Unigold) were available. Serial-algorithm was used in all the laboratories. In 31 (70.4%) of them external quality control specimens were not used. Twenty two (50.0%) of the laboratories reported frequent shortage of reagents. All (100%) were able to identify negative specimens distributed. Positive proficiency panel samples were identified in 37 (94.8%) of the 39 laboratories. There was 98.3% agreement at a screening level between the sites and the central laboratory. Rate of discrepancy between screening and confirmatory assays was found to be 3.0% and 2.1% at the sites and at central laboratory, respectively., Conclusion: The test kits showed a good performance at the point of services in the field sites. However, continuous assessment of HIV test kits at the point of service and training of professionals on newly arrived techniques are recommended to have effective testing performance with acceptable sensitive and specific testing algorithm. Effective quality assurance program should be in place to support programs such as VCT, prevention of mother-to-child-transmission and antiretroviral therapy.

Introduction: Infection is the most serious complication of diabetes and recognized as causing significant morbidity and mortality., Objectives: To assess the prevalence and pattern of infection, determine the causative organisms., Methods: A cross sectional study involved 179 diabetic patients admitted to the Department of Internal Medicine of Tikur Anbessa Specialized University Hospital performed between Nov. 2000 - Nov 2002, clinical data were documented, a total of 114 clinical specimens (40 pus, 23 blood and 51 urine) were submitted to bacteriology laboratory for bacterial isolation and identification., Results: The mean +/- SD fasting blood sugar was 246.0 +/- 153.3 mg/dl. Fifty eight (32.4%) of patients were admitted in state of diabetic ketoacidosis. The prevalence of infection in diabetic patients was 44%. Diabetic foot infection (35%) was the commonest, followed by pulmonary tuberculosis 22%, urinary tract infection (14%), pneumonia (12.8%), skin and subcutaneous infection (12.8%). About 30% of bacterial isolates were Staphylococcus aureus followed by Klebsiella pnumeniae (23.4%), Escherichia coli (19%) and Pseduomonas spp (15%). The rest of the isolates were less frequent and ranged between 2 to 6.4%. More than one bacterial species were also isolated from 10% of the positive cultures. Main causes of deaths in hospitalized diabetic patients were cardiovascular diseases (6/15), end stage renal disease (3/15) and sepsis (2/15)., Conclusion: Infections were the leading cause of morbidity while cardiovascular diseases were the leading cause of mortality. Diabetic foot ulcers were the major cause of infection followed by tuberculosis, skin and subcutaneous infections and pneumonia. The prevalence of tuberculosis in diabetic patient is increasing; S. aureus from wound infection and E. colifrom urinary tract infection were the common pathogens.

We studied the use of dried spots of bodily fluids (plasma, whole blood, and mother's milk) on filter paper as a means of sample collection and storage for human immunodeficiency virus type 1 (HIV-1) viral load testing under stringent field conditions. Plasma placed directly in lysis buffer, which is customarily used for viral load assays, was used for comparison in all our experiments. Utilizing reconstruction experiments, we demonstrate no statistical differences between viral loads determined for plasma and mother's milk spotted on filter paper and those for the same fluids placed directly in lysis buffer. We found that the addition of whole blood directly to lysis buffer was unreliable and could not be considered a feasible option. However, viral load measurements for whole blood spotted onto filter paper correlated with plasma viral load values for both filter spots and lysis buffer (Pearson correlation coefficients, 0.7706 and 0.8155, respectively). In conclusion, dried spots of plasma, whole blood, or mother's milk provide a feasible means for the collection, storage, and shipment of samples for subsequent viral load measurement and monitoring. Virus material spotted and dried on filter paper is a good inexpensive alternative for collecting patient material to monitor the HIV-1 viral load. Measuring the HIV-1 burden from whole blood dried on filter paper provides a suitable alternative for low-technology settings with limited access to refrigeration, as can be found in sub-Saharan Africa.

There is limited information on the biology and pathogenesis of Leishmania aethiopica, causative agent of cutaneous leishmaniasis (CL) in Ethiopia. In this study we have identified and characterized two cathepsin L-like cysteine protease genes, Laecpa and Laecpb, from L. aethiopica. The predicted amino acid sequence of Laecpa and Laecpb is more than 75% identical with homologous cathepsin L-like cysteine protease genes of other Leishmania species and less than 50% identical with human cathepsin L. Laecpa is expressed predominantly in the stationary, and to a lower level, during the amastigote stage while Laecpb is specifically expressed in the stationary stage of L. aethiopica development. Phylogenetic analysis showed that the two genes are grouped into separate clades which are the result of gene duplication. The isolation of these genes will be useful in developing Leishmania species specific diagnostics for molecular epidemiological studies and serves as a first step to study the role of cysteine proteases in L. aethiopica pathogenesis.

Forty-nine samples with known C2V3 sequences were used for the evaluation of an env-based molecular beacon assay to distinguish between the two genetic subclusters C and C' which characterize the HIV-1 epidemic in Ethiopia. Two subcluster C and two subcluster C' beacons targeting two different loci in the C2V3 region were developed. Using a three beacon-based (2C and 1C'=C prime), isothermal amplification assay, concordance with DNA sequencing was achieved for 43 (87.8%) samples. Sensitivity was 81.8% and specificity 97.4% for subcluster C beacons. For the subcluster C' beacon, a sensitivity of 97% and a specificity of 87.5% was achieved. Five samples were ambiguous by sequencing of which two samples were subcluster C' by the beacon assay and one subcluster C. Two of the samples remained ambiguous with different beacon-pair combinations as well. From samples with a clear C or C' phylogeny by sequencing, three were undetected by the first-line beacon genotyping assay. Genotype ambiguity was resolved in the three samples using beacon pair combinations restricted to each targeted locus. The beacons were evaluated further in a panel including all HIV-1 subtypes. Four of five subtype C isolates were identified correctly, and no cross-reactivity was observed with other subtypes.

The potential medical transmission of HIV through unsafe medical injections was evaluated in 16 rural health institutions in Ethiopia. Most institutions reported re-using disposable needle/syringes, and 12% of observed injections were given with used, disposable syringes prepared for re-use. Analysis of used needle flushes showed no HIV RNA; however, the sensitivity of our method was limited. Despite the re-use of disposable needles, medical injection practices are not likely to contribute significantly to HIV transmission in this region.

The objective of this investigation was to assess the prevalence of dermatophytoses in children in a geographically restricted area in the Ethiopian countryside, and to determine the aetiological agents of these infections. Demographical and clinical-dermatological data were collected from all children 4-15 years of age on Tulugudu Island, Southern Ethiopia. Mycological specimens were taken and species identification determined through morphological observations and biochemical tests, complemented with sequencing of rDNA ITS2 region when necessary. Of 171 children, 96% shared combs, 85% shared beds and 97% had animal contact. Family size was > 5 persons in 50% of the test subjects and prevalence of tinea capitis was elevated in this group (P < 0.005). Dermatophytoses were clinically diagnosed in 136 cases (79.5%). Tinea capitis (T. capitis) was the most common manifestation with 104 cases (76.5%). T. capitis was combined with dermatophytic infections at other sites in 19 cases. Tinea faciae and Tinea corporis were found in four and two cases, respectively, and pediculosis capitis was diagnosed in 2.9% of the test subjects. Of 135 samples from hair (n = 112), skin (n = 19) and finger-nail (n = 4), 74.1% were microscopy-positive for dermatophytes, 73% were positive in culture, giving an overall prevalence of dermatophytoses in 57.3% of all children examined. Trichophyton violaceum was identified in 80.6% of cultures, Trichophyton verrucosum in 16.3% and Trichophyton tonsurans in 2.0%. One isolate was identified as a white variant of T. violaceum. Tinea capitis was highly prevalent in children on Tulugudu Island, Southern Ethiopia. The anthropophilic species T. violaceum dominated as an aetiological agent. Zoophilic dermatophytes were relatively rarely isolated from clinical specimens, despite the children's frequent contact with animals.

The herpes simplex virus type 2 (HSV-2) and human immunodeficiency virus (HIV) epidemics are believed to fuel each other, especially in sub-Saharan countries. In Ethiopia during 1997-2002, a retrospective study was conducted to examine risk factors for infection and transmission of HSV-2, in a cohort of 1612 factory workers. Prevalence of HSV-2 seropositivity at enrollment was 40.9%, and incidence of seroconversion was 1.8 seroconversions/100 person-years (PY), which decreased over time. Independent risk factors for seropositivity were having an HSV-2-seropositive partner, female sex, HIV antibodies, positive Treponema pallidum particle agglutination assay result, older age, low education level, and orthodox religion. These same factors were independent risk factors for HSV-2 seroconversion, with the exception of the latter 3. Most HSV-2-infected persons did not report symptoms. Among 41 monogamous HSV-2-serodiscordant heterosexual couples, incidence of HSV-2 seroconversion was 20.75 seroconversions/100 PY for women and 4.93 seroconversions/100 PY for men. The high burden of both HSV-2 and HIV infection in Ethiopia warrants stringent control measures.

A total of 300 gastric biopsy samples and 50 Helicobacter pylori isolates were collected from Ethiopian adult dyspeptic patients. The vacA and cagA genes were detected in 90 and 79% of biopsy specimens, respectively, and in 100 and 87% of clinical isolates, respectively. Both genes were detected in 84% of the gastric biopsy samples and in 87% of the clinical isolates. Among vacA genotypes, the s1/m1 genotype was the most common in gastric biopsy samples (48%). The vacA and cagA positive H. pylori strains were detected to a higher degree in patients with chronic active gastritis (71%) than patients with other histopathological findings (29%) (P < 0.05).

Rifampin is a major drug used in the treatment of tuberculosis infections, and increasing rifampin resistance represents a worldwide clinical problem. Resistance to rifampin is caused by mutations in the rpoB gene, encoding the beta-subunit of RNA polymerase. We examined the effect of three different rpoB mutations on the fitness of Mycobacterium tuberculosis. Rifampin-resistant mutants were isolated from a virulent clinical isolate of M. tuberculosis (strain Harlingen) in vitro at a mutation frequency of 2.3 x 10(-8). Mutations in the rpoB gene were identified, and the growth rates of three defined mutants were measured by competition with the susceptible parent strain in laboratory medium and by single cultures in a macrophage cell line and in laboratory medium. All of the mutants showed a decreased growth rate in the three assays. The relative fitness of the mutants varied between 0.29 and 0.96 (that of the susceptible strain was set to 1.0) depending on the specific mutant and assay system. Unexpectedly, the relative fitness ranking of the mutants differed between the different assays. In conclusion, rifampin resistance is associated with a cost that is conditional.

Presently, there is no published information on the antimicrobial susceptibility pattern of H. pylori strains in Ethiopia to guide the choice of drug for therapy. Therefore, it is becoming clinically relevant to test the in vitro susceptibility of H. pylori clinical isolates prior to treating patients. Susceptibility testing was performed on 50 clinical H. pylori isolates obtained from adult dyspeptic patients referred to the gastrointestinal (GI) Clinic of Tikur Anbassa University Hospital. Five antibiotics were evaluated, by using the Episolmeter test (E-test). The antibiogram of 50 H. pylori clinical isolates showed that all strains were sensitive to clarithromycin, erythromycin and tetracycline, while 38/50 (76%) and 3/50 (6%) of the strains were resistant to metronidazole and amoxicillin, respectively. Infection by metronidazole or amoxicillin resistant H. pylori is an important factor leading to treatment failure. Testing of all H. pylori clinical isolates to metronidazole and amoxicillin is recommended. If it is not possible to perform susceptibility tests on each clinical isolate, a program to survey the prevalence of resistance should be implemented in a given area or population. When treatment of H. pylori infection is indicated in dyspeptic patients, the potential availability, simplicity of use, safety and low cost of the antimicrobial agents have to be taken into account.

A gag-based molecular beacon assay utilizing real-time nucleic acid sequence-based amplification technology has been developed to differentiate between the two genetic subclusters of human immunodeficiency virus type 1 (HIV-1) subtype C (C and C') circulating in Ethiopia. Of 41 samples, 36 could be classified as C or C' by sequencing of the gag gene. All 36 isolates were correctly identified by the gag beacon test. Three isolates with genomes that were recombinant in gag were unambiguously typed as belonging to the C' subcluster. Further analysis revealed that these contained the most sequence homology with a reference subcluster C' sequence in the target region of the beacon and hence were correct for the analyzed region. For one sample, sequencing and gag molecular beacon results did not match, while another isolate could not be detected at all by the beacon assay. Overall, high levels of sensitivity and specificity were achieved for both beacons (90.5% sensitivity and 100% specificity for the C beacon and 100% sensitivity and 95.2% specificity for the C' beacon). The availability of a diagnostic test which can quickly and reliably discriminate between C and C' HIV-1 infections in Ethiopia is an important first step toward studying their respective biological characteristics. As the assay is specific to the Ethiopian HIV-1 subtype C epidemic, it will contribute to characterizing the circulating viruses in this population, thereby generating the information necessary for the development of a potential efficacious HIV-1 vaccine appropriate for the Ethiopian context.

Individuals infected with human immunodeficiency virus type 1 (HIV-1) subtype C infrequently harbour X4 viruses. We studied R5 and X4 biological clones generated from HIV-1 subtype C-infected individuals. All subtype C R5 viruses demonstrated slower profiles of replication on CD4(+) lymphocytes in comparison to subtype B viruses, whereas subtype C X4 viruses replicated with comparable efficiency to subtype B X4 viruses. No differences were identified in CC or CXC chemokine inhibitions (RANTES and SDF-1alpha, respectively) between subtype C and subtype B viruses. Immature dendritic cells were shown in coculture experiments to similarly enhance the infection of subtype C and subtype B R5 as well as X4 viruses. By amino acid sequence analysis, we showed that the R5 and X4 subtype C gp120 envelope gene alterations were similar to those for a switching subtype B virus, specifically with respect to the V3 charge and envelope N-linked glycosylation patterns. By phylogenetic analysis, we showed that one patient was infected with HIV-1 C' and the other was infected with HIV-1 C" and that one of the patients harbored a virus that was a recombinant in the gp120 env gene between an R5 and an X4 virus, with the resultant virus being R5. No differences were identified between the long terminal repeat regions of the subtype C R5 and X4 biological clones. These results indicate that even though R5 subtype C viruses are restrictive for virus replication, the R5-to-X4 phenotype switch can occur and does so in a manner similar to that of subtype B viruses.

The magnitude and complexity of the HIV-1 genetic diversity are major challenges for vaccine development. Investigation of the genotypes circulating in areas of high incidence, as well as their interactions, will be a milestone in the development of an efficacious vaccine. Because HIV-1 subtype C (HIV-1C) is responsible for most of the 36 million infections worldwide we investigated the HIV-1C strains circulating in Ethiopia in a retrospective, cross-sectional study. Serum samples from HIV-1-positive individuals were collected in seven Ethiopian cities and towns. Nucleotide sequences of the gag, pol, and env genes were analyzed. We performed phylogenetic analysis by the neighbor-joining and maximum-likelihood methods with sequences from 30 isolates, and we determined recombination by the bootscanning method as implemented in the SIMPLOT program. Sequence analyses of a 2600-nucleotide fragment (including the gag gene, the protease, and the 5' half of reverse transcriptase of the pol gene) and the corresponding V1V2/C2V3 envelope regions confirmed that two distinct HIV-1C genotypes (C' and C") are cocirculating in Ethiopia, as shown previously by the analysis of the C2V3 envelope region. We have identified intrasubtype recombination between the two HIV-1C genotypes, C' and C", with 6 of the 30 (20%) analyzed viruses being recombinants. The C' sequences were phylogenetically linked to the fast spreading viruses in India and southern Africa. Furthermore, all the recombinant viruses shared the C' V1V3 region of the envelope, suggesting that the prevalence of viruses with the C' envelope is increasing compared to the C" envelope. The possibility that viruses with a C' envelope have a biological advantage over the viruses with a C" envelope should be further investigated in biological and epidemiological studies.

Objective: To investigate the susceptibility of clinical isolates of gram-negative bacteria to chlorhexidine gluconate., Design: Prospective laboratory study., Setting: Tikur Anbessa Hospital, Addis Ababa, Ethiopia., Subjects: Clinical specimens from 443 hospital patients., Main Outcome Measures: Significant number of gram negative bacteria were not inhibited by chlorhexidine gluconate (0.02-0.05%) used for antisepsis., Results: Four hundred and forty three strains of gram-negative bacteria were isolated from Tikur Anbessa Hospital patients. Escherichia coli (31.6%) and Klebsiella pneumoniae (23%) were the most frequently isolated bacteria followed by Proteus species (13.3%), Pseudomonas species (9.2%), and Citrobacter species (6.1%). Each organism was tested to chlorhexidine gluconate (CHG), minimum inhibitory concentration (MIC) ranging from 0.0001% to 1%w/v. All Salmonella species and E. coli were inhibited by CHG, MIC < or = 0.01%. Twenty nine per cent of Acinetobacter, 28% of K. pneumoniae and Enterobacter species and 19-25% of Pseudomonas, Proteus and Providencia species were only inhibited at high concentrations of CHG (> or = 0.1%)., Conclusion: Our results showed that a significant number of the gram-negative bacterial isolates were not inhibited by CHG at the concentration used for disinfection of wounds or instruments (MIC 0.02-0.05% w/v). It is therefore important to select appropriate concentration of this disinfectant and rationally use it for disinfection and hospital hygiene. Continuing follow up and surveillance is also needed to detect resistant bacteria to chlorhexidine or other disinfectants in time.

The synthesis of capsular polysaccharide by Klebsiella pneumoniae K1 was investigated in a minimal salts medium by continuous culture. The organism produced larger amounts of polysaccharide under nitrogen-limited conditions than under carbon-limited conditions. The synthesis of polysaccharide was dependent not only on the availability of excess carbon, but also on growth rate. The rate of polysaccharide synthesis was greatest at low dilutions, low temperature (30 degrees C) and at neutral pH. Prolonged growth in nitrogen-limited culture resulted in the development of non-mucoid variants, possibly due to a selective growth advantage over mucoid cells. The non-mucoid isolate was more susceptible to some bacteriophages, possibly due to the reduction nor absence of capsular polysaccharide.

One thousand pharyngeal swab specimens were processed for aerobic culture to determine the carriage rate of Gram-negative bacilli (GNB). The isolates were identified and their sensitivity determined to 11 antibacterial drugs following standard techniques. Similar pharyngeal carriage rates of GNB were found among the various groups of healthy subjects. Patients had higher colonization rates (27%) than healthy subjects (16%). The increase in prevalence of GNB seemed to be associated with underlying diseases and duration of hospitalization. Klebsiella (36%) was the most frequent genus amongst the 215 isolates of GNB followed by pseudomonas (13%), enterobacter (13%) and acinetobacter (10%). Others were less frequently isolated. Over 70% of all isolates were resistant to ampicillin (79%) and carbenicillin (72%); 55, 45 and 43% were resistant to cephalothin, tetracycline and streptomycin, respectively. The great majority of the strains were sensitive to the remaining six drugs. The hospital isolates were more resistant than the non-hospital isolates to most drugs tested. The hospital strains were also more often multiply resistant (89%) than the non-hospital strains (60%). Sixty-five different resistance antibiograms of 1-10 drugs were observed among 191 strains. More varied types of antibiograms were observed among hospital strains. The high frequency of multiple drug resistance of the isolates is an indication of the extensive use of antibacterial drugs, indicating the need for a policy for judicious use of drugs.