Genetic characterization of antimicrobial resistance of Shigella flexneri 1c isolates from patients in Egypt and Pakistan.

BACKGROUND: Shigella flexneri serotype 1c emerged as a critical isolate from children in Egypt and Pakistan. The pattern of antimicrobial susceptibility (AMS) and resistance genes of this serotype have yet to be characterized. FINDINGS: Sixty nine S. flexneri 1c isolates isolates were identified from both Egypt (n-46) and Pakistan (n = 23) and tested for AMS by disk diffusion method and minimal inhibitory concentrations were also determined. Isolates were genotyped by pulsed field gel electrophoresis (PFGE) and five relevant resistance genes (bla(TEM), bla(SHV), bla(OXA), sulI and sulII) were detected by polymerase chain reaction (PCR) and confirmed by DNA sequencing. High resistance was observed in all isolates for ampicillin (AM >96%); trimethoprim-sulphamethoxazole and tetracycline (>88%). Most AM-resistant isolates from Egypt (70%) harbored bla(TEM) resistance, while 52% of isolates from Pakistan expressed bla(OXA). All isolates were closely related by PFGE, irrespective of source or time of collection. The sulII gene was present in 100% of isolates from pediatric cases in Egypt, 65% of Pakistan isolates, and 53% of isolates from older Egyptian patients. CONCLUSIONS: While different Shigella serotypes gathered in specific genotypic groups, 1c serotype isolates formed multiple clusters. Although AMS was considerably high to most commonly used drugs, genetic determinants were variable between countries over time. The data stress the need for a more careful selection of antibiotics in the treatment of shigellosis.

A simple technique for long-term preservation of leptospires.

The viability of six serovars of Leptospira spp. was studied after long storage at -70°C. The bacteria were either preserved in Ellinghausen-McCullough-Johnson-Harris (EMJH) liquid growth medium or in sheep blood added as a cryoprotectant. The viability of the strains was observed on a monthly basis by dark-ground microscopy over a period of 20 months at -70°C. Addition of sheep blood was not significantly advantageous, since leptospires that were stored in EMJH showed a slight increase in number after recovery. The results suggest a very simple and useful technique for long-term preservation of such Leptospira.

Concurrent infections in acute febrile illness patients in Egypt.

We report the occurrence of concurrent infections with multiple acute febrile illness (AFI) pathogens during an ongoing prospective laboratory-based surveillance in four infectious disease hospitals in urban and rural areas of Egypt from June 2005 to August 2006. Patients were screened for Leptospira, Rickettsia typhi, Brucella, or Salmonella enterica serogroup Typhi by various methods including serology, culture, and PCR. One hundred eighty-seven of 1,510 patients (12.4%) evaluated had supporting evidence for the presence of co-infections; 20 (1%) of these patients had 2 or more pathogens based upon confirmatory 4-fold rise in antibody titer, culture, and/or PCR. Most coinfected patients lived or worked in rural agricultural areas. The high coinfection rates suggest that defining the etiologies of AFI is imperative in guiding proper disease treatment, prevention, and control strategies in Egypt.

Brucellosis as a cause of acute febrile illness in Egypt.

To develop better estimates of brucellosis incidence, we conducted population-based surveillance for acute febrile illness (AFI) in Fayoum governorate (population 2347249), Egypt during two summer periods (2002 and 2003). All hospitals and a representative sample of community healthcare providers were included. AFI patients without obvious etiology were tested for brucellosis by culture and serology. Incidence estimates were calculated adjusting for sampling methodology and study period. Of 4490 AFI patients enrolled, 321 (7%) met the brucellosis case definition. The estimated annual incidence of brucellosis per 100000 population was 64 and 70 in 2002 and 2003, respectively. The median age of brucellosis patients was 26 years and 70% were male; 53% were initially diagnosed as typhoid fever. Close contact with animals and consumption of unpasteurized milk products were associated with brucellosis. The high incidence of brucellosis in Fayoum highlights its public health importance, and the need to implement prevention strategies in humans and animals.

Occurrence of ampicillin-resistant Enterococcus faecium carrying esp gene in pet animals: An upcoming threat for pet lovers.

OBJECTIVES: This study was carried out to investigate oral colonisation by Enterococcus faecalis and Enterococcus faecium in pet dogs and cats, with special reference to antibiotic resistance. METHODS: Oral swabs were collected from 63 pet dogs and 57 pet cats with no known history of hospitalisation. All samples were enriched in Kenner Fecal (KF) broth before being cultured on KF agar to isolate enterococci. E. faecalis and E. faecium were identified by biochemical and molecular techniques. Antimicrobial resistance was determined by the disk diffusion method, and ampicillin-resistant strains were further examined by PCR to detect the esp gene. RESULTS: Oral prevalence rates of E. faecalis among pet dogs and cats were 3.2% and 5.3%, respectively, whilst those for E. faecium were 22.2% and 15.8%, respectively. None of the isolated enterococci were resistant to vancomycin. However, ampicillin-resistant E. faecium (AREfm) was detected in the examined dogs and cats at rates of 14.3% and 5.3%, respectively. Moreover, among the isolated enterococci, six isolates showed multidrug resistance (all AREfm). Whilst the esp gene was detected in only two of nine canine AREfm isolates (multidrug-resistant strains), none of feline AREfm isolates harboured esp. CONCLUSIONS: The occurrence of AREfm and the esp gene among oral isolates from pet dogs and cats represents a great public health hazard for pet owners and highlights possible zoonotic transmission of such a nosocomial pathogen outside healthcare facilities.

Retrospective serosurvey of leptospirosis among patients with acute febrile illness and hepatitis in Egypt.

The epidemiologic status of leptospirosis in Egypt has not been well defined because of difficulties in disease diagnosis. A retrospective study was conducted to detect leptospiral antibodies among undiagnosed acute febrile illness (AFI) and hepatitis cases. Approximately 16% of both AFI (141/886) and acute hepatitis (63/392) cases showed seroreactivity to Leptospira IgM by ELISA and microscopic agglutination test (MAT). Canicola, Djasiman, Grippotyphosa, Pyrogenes, Icterohemorrhagiae, and Pomona were the most commonly reactive serovars among patients with AFI. Djasiman, Grippotyphosa and Icterohemorrhagiae were the most reactive among patients with acute hepatitis. This study represents the first systematic report of Leptospira associated with patients with AFI and hepatitis in Egypt. Physicians need to have increased awareness about the importance of leptospirosis in the differential diagnosis of AFI and acute hepatitis in Egypt. In addition, laboratory capacity should be developed at fever hospitals to diagnose leptospirosis.

Rapid enzyme-linked immunosorbent assay for the diagnosis of human brucellosis in surveillance and clinical settings in Egypt.

OBJECTIVE: To optimize and standardize an enzyme-linked immunosorbent assay (ELISA) for rapid diagnosis of human brucellosis in clinical cases identified during a surveillance study for acute febrile illness (AFI). METHODS: Serum samples from patients presenting with AFI at 13 fever hospitals across Egypt between 1999 and 2003 were kept frozen at NAMRU-3 and used in this study. The assay was evaluated in 5 subject groups: brucellosis cases confirmed by blood culture (group I, n=202) 87% positive by standard tube agglutination test (TA), brucellosis cases exclusively confirmed by TA (group II, n=218), blood cultures from AFI cases positive for bacterial species other than Brucella (group III, n=103), AFI cases with unexplained etiologies (group IV, n=654), and healthy volunteers (group V, n=50). All members of groups III-V were negative for brucellosis by TA. RESULTS: Sensitivity and specificity of ELISA for total specific antibodies were >=96% versus 87% for TA as compared to microbial culture, the current gold standard method for Brucella identification. Assessment of Brucella antibody classes by ELISA in random subsets of the 5 groups showed significantly high (p>0.001) levels of anti Brucella IgG (>=81%) and IgM (>=90%) in groups I and II only. CONCLUSION: The obtained sensitivity and specificity results indicate that our ELISA is more suitable for AFI surveillance and clinical settings than blood culture and TA. The developed assay is also cost-effective, easier to use, faster, and the coated plates can be stocked for at least 8 months, providing a potential for field use and automation.

Trends of multiple-drug resistance among Salmonella serotype Typhi isolates during a 14-year period in Egypt.

A total of 853 isolates of Salmonella serotype Typhi recovered from patients with typhoid fever who were admitted to a major infectious disease hospital in Cairo, Egypt, from 1987 through 2000 underwent antibiotic susceptibility testing to determine multiple-drug resistance. The observed resurgence of chloramphenicol susceptibility (P=.002) may suggest reuse of this drug for the treatment of typhoid fever in Egypt.

Phenotypic diversity of enterotoxigenic Escherichia coli (ETEC) isolated from cases of travelers' diarrhea in Kenya.

BACKGROUND: The aim of this study was to characterize phenotypically enterotoxins, colonization factors (CFs) and the antibiotic susceptibility of enterotoxigenic Escherichia coli (ETEC) strains isolated from cases of acute diarrhea that occurred in Europeans traveling to resorts in Mombasa, Kenya; this information is critical for the development of vaccines and empirical treatment. METHODS: Over a 1-year period from 1996 to 1997, five E. coli-like colonies were obtained from each of 463 cases with acute diarrhea. These strains were characterized for enterotoxins using GM-1 ELISA, for CFs using a dot-blot assay, and for antibiotic susceptibility using antibiotic disks. RESULTS: Of 164 strains characterized for ETEC phenotype, 30 (18%) expressed heat-labile toxin (LT) only, 83 (51%) heat-stable toxin (ST) only, and 51 (31%) both LT and ST. Analysis for CF expression demonstrated that 107 (65%) of the strains were positive for CFs, including CFA/IV (46%), CFA/II (35%), and CFA/I (5%), while less than 4% expressed less common CFs. All ETEC strains tested were resistant to erythromycin and sensitive to ceftriaxone. Over one-third of the strains were resistant to sulfamethoxazole-trimethoprim or tetracycline. Six strains were resistant to nalidixic acid; none of these were resistant to ciprofloxacin. CONCLUSIONS: Cumulatively, our findings indicate that ETEC in this region comprises a highly diverse group of bacterial enteropathogens, and that the development of prophylactic agents against ETEC faces major challenges because of this diversity.

Serological evidence of flaviviruses infection among acute febrile illness patients in Afghanistan.

INTRODUCTION: Current published reports on the causative agents of acute febrile illness (AFI) in Afghanistan are scarce, and the burden of disease due to flaviviruses is unknown. METHODOLOGY: A hospital-based surveillance study for AFI was established in 2008 through 2010 to determine the seroepidemiology of West Nile virus (WNV), tick-borne encephalitis virus (TBEV) and dengue viruses (DENV) using commercial ELISA kits. Due to major logistical challenges, only acute sera were collected. RESULTS: Serological analysis for IgG were as follows: WNV 30.4% (277/913); TBEV 23.4% (214/913); DENV 19.7% (180/913). Single positive IgG reactions for WNV, TBEV and DENV were noted in 11% (100/913), 7.2% (66/913), and 5% (47/913), respectively. Reactivity for all three screened flaviviruses was detected in 44.5% (406/913) of sera. IgM positivity was uncommon, with only 0.5% (5/913), 2.2% (20/913) and 2.6% (8/312) of samples positive for WNV, TBEV, and DENV, respectively. Serological findings were confirmed in random positive samples by neutralization assay. CONCLUSIONS: These serological results suggest circulation of WNV, TBEV, and DENV within Afghanistan, with evidence of current or prior infection noted in a significant proportion of patients seeking care for AFI. Obtaining additional information on the prevalence of these and other causes of AFI is paramount for improving the distribution of available limited syndromic treatment and improving the existing health protection policy in Afghanistan.

Heterogeneity of Campylobacter species isolated from serial stool specimens of Egyptian children using pulsed field gel electrophoresis.

BACKGROUND: The genus Campylobacter spp. is a common cause of human acute bacterial enteritis and travellers' diarrhoea worldwide. OBJECTIVE: To determine whether multiple serial isolations of Campylobacter spp., when obtained from a single child, represented the same or a different organism. METHODS: In a birth cohort study conducted in Egypt, numerous children showed serial isolations of Campylobacter spp. Of these, 13 children were selected from different households based on the successive isolation of six or more Campylobacter isolates from stool samples. RESULTS: Eighty isolates were recovered and identified as either Campylobacter coli (n = 25) or Campylobacter jejuni (n = 55). Pulsed-field gel electrophoresis (PFGE) revealed the presence of 38 unique C. jejuni and 24 C. coli profiles at a similarity level of ≥ 90%. Although no serially-identical isolates were detected in six children, others demonstrated at least one identical couple of isolates; all identified serially between one to six weeks. Two children demonstrated > 80% similar couples of isolates that appeared seven months apart. PFGE could be a useful tool for differentiating reinfection, relapse and convalescent excretion phases. CONCLUSION: Our data suggest that Campylobacter infection in children is a complex process; children are exposed to multiple species in endemic environments and strains of the same bacterium appear to be shed serially between one to six weeks after the first exposure. Isolates that persisted for longer periods were relatively less similar, as shown from the results of this study.

Building military influenza surveillance capacity in West Africa.

Militaries are especially susceptible to operationally important outbreaks of acute respiratory infections such as pandemic and seasonal influenza. In addition, militaries play important roles for State Parties working to meet International Health Regulations 2005, particularly in developing countries. In 2009, the U.S. Naval Medical Research Unit No. 3 joined with the Noguchi Memorial Institute for Medical Research and the armed forces of Ghana, Burkina Faso, and Côte d'Ivoire to create or improve influenza surveillance capacities within the militaries. This article describes the process undertaken to achieve this goal. In the Ghana Armed Forces, influenza surveillance for outpatients was instituted at seven medical stations throughout the country and for inpatients at the tertiary referral hospital in Accra. As a result, military sites now contribute around half of the influenza cases detected in Ghana and reported weekly to the World Health Organization. Samples were also collected by the militaries of Côte d'Ivoire and Burkina Faso, although political instability slowed progress. This effort is part of an ongoing strategy to build influenza surveillance capacity within West African militaries in support of military services, global outbreak investigations, International Health Regulations-2005, and the development of country-specific pandemic preparedness plans.

Characterization of Neisseria meningitidis isolates from Egypt using multilocus sequence typing.

To characterize Neisseria meningitidis isolates collected from cerebrospinal fluid of meningitis cases in Egypt (1998-2003) as part of surveillance studies, 67 isolates were serogrouped, tested for antibiotic sensitivity and analyzed using multilocus sequence typing (MLST). Results show that isolates expressing serogroup B (50.7%) and serogroup A (34.3%) antigens were predominant in Egypt during the surveillance period, possibly due to suppression of other serogroups by meningococcal vaccines in current use. Intermediate resistance to penicillin was observed in 71% of the isolates, suggesting a need for physicians to shift to third-generation cephalosporins during the empirical treatment of infection. Recurrent lineages of N. meningitidis in Egypt appear to originate from Europe and other Middle Eastern countries. Of 19 sequence types detected, five were unique to Africa and 10 were not observed previously in the MLST database. The information obtained illustrates the changing dynamics of meningitis after vaccine introduction in Egypt.

Antimicrobial resistance in pathogens causing nosocomial bloodstream infections in university hospitals in Egypt.

BACKGROUND: Nosocomial bloodstream infections (BSIs) and antimicrobial resistance (AMR) are worldwide health care problems causing substantial patient morbidity and mortality. This study was conducted to identify bacterial pathogens isolated from nosocomial BSIs and determine their AMR patterns. METHODS: An active surveillance program for BSIs was conducted in intensive care units in 3 large university hospitals in Egypt between September 1, 2006, and June 30, 2007. Infection prevention and control teams and link nurses in collaboration with intensive care physicians were looking actively to identify patients who acquired BSIs based on Centers for Disease Control and Prevention standard case definitions. Blood cultures were obtained from patients with suspected BSIs and processed to isolate bacteria and test their antimicrobial resistance. RESULTS: During the 10-month active surveillance period, a total of 600 pathogens were isolated from blood cultures of 1,575 patients (38%). Of these 600 isolates, 386 (66%) were gram-negative, 178 (30%) were gram-positive, and 24 (4%) were budding yeasts. The gram-negative organisms included 162 (27%) Klebsiella pneumoniae and 23 (3.8%) Escherichia coli. Extended-spectrum ß-lactamase enzymes were detected in 79% of the K pneumoniae isolates and 39% of the E coli isolates. Methicillin-resistant Staphylococcus aureus accounted for 60% of S aureus infections. CONCLUSIONS: High rates of ß-lactamase resistance and methicillin-resistant S aureus were found in the 3 Egyptian university hospitals studied. This study highlights the need for strengthening infection prevention and control programs, monitoring AMR at each facility, and developing policies for antibiotic use.

Cross-species surveillance of Leptospira in domestic and peri-domestic animals in Mahalla City, Gharbeya Governorate, Egypt.

A survey of 179 animals (black rats, dogs, sheep, buffaloes, cattle, donkeys, weasels, and cats) for Leptospira infection was conducted in Mahalla City (Lower Egypt). Blood, urine, and kidney were collected and tested by culture, microscopic agglutination test (MAT), and/or polymerase chain reaction (PCR). Among rats, 26% were positive by PCR, including 7% that were also positive by culture for L. interrogans serovars Grippotyphosa, Pyrogenes, and Icterohaemorrhagiae. L. borpetersenii serovar Polonica was isolated for the first time in Egypt in three rats. MAT titers ≥ 1:800 were observed in 11% of rats and 12% of dogs. L. interrogans serovar Grippotyphosa was detected in one cat. Sheep and donkeys were negative for leptospirosis by all methods. Buffaloes and cattle were seropositive in 20% and 44% of animals, respectively. Data indicate that several pathogenic serovars are circulating in the animals, which may pose exposure risks and account for high rates of acute febrile illness.

Enteric fever in Mediterranean north Africa.

Typhoid fever is endemic in the Mediterranean North African countries (Morocco, Algeria, Tunisia, Libya, and Egypt) with an estimated incidence of 10-100 cases per 100,000 persons. Outbreaks caused by Salmonella enterica serovar Typhi are common and mainly due to the consumption of untreated or sewage-contaminated water. Salmonella enterica Paratyphi B is more commonly involved in nosocomial cases of enteric fever in North Africa than expected and leads to high mortality rates among infants with congenital anomalies. Prevalence among travellers returning from this region is low, with an estimate of less than one per 100,000. Although multidrug resistant strains of Salmonella Typhi and Paratyphi are prevalent in this region, the re-emergence of chloramphenicol- and ampicillin-susceptible strains has been observed. In order to better understand the epidemiology of enteric fever in the Mediterranean North African region, population-based studies are needed. These will assist the health authorities in the region in preventing and controlling this important disease.

An effective venipuncture technique and normal serum biochemistry parameters of the captive fat-tailed jird (Pachyuromys duprasi).

Thirty-nine captively reared fat-tailed jirds (Pachyuromys duprasi) were enrolled in a minimally invasive study to determine an effective venipuncture technique and establish normal serum biochemistry parameters. A jugular venipuncture technique using chemical restraint (ketamine, 30 mg/kg; xylazine, 6 mg/kg; acepromazine, 1 mg/kg) administered intraperitoneally was safe and consistently yielded at least 0.3 mL of blood. Of the biochemical indicators measured (glucose, total protein, albumin, globulin, alkaline phosphatase, alanine transferase, total bilirubin, amylase, BUN, creatinine, calcium, phosphorous, sodium and potassium), amylase and glucose levels differed significantly between male and female fat-tailed jirds.

Antimicrobial susceptibility and serotype distribution of Streptococcus pneumoniae causing meningitis in Egypt, 1998-2003.

OBJECTIVES: To determine the antimicrobial susceptibility and serotype distribution of 205 isolates of Streptococcus pneumoniae, collected from the CSF of meningitis patients identified between 1998-2003, during sentinel meningitis surveillance in Egypt. METHODS: Antimicrobial susceptibility was evaluated against six antibiotics using disc diffusion and Etest methods. Serotyping was performed by latex agglutination and the Quellung test. RESULTS: Forty-nine percent of all isolates were found to be non-susceptible to penicillin (46% intermediate, MIC range 0.12-1.0 mg/L; 3% resistant, MIC = 2.0 mg/L), and 6% of the isolates were non-susceptible to ceftriaxone (5% intermediate, MIC = 1.0 mg/L; 1.3% resistant, MIC >/= 2 mg/L). Resistance rates for tetracycline and trimethoprim/sulfamethoxazole were high (52 and 59.7%, respectively), but those for erythromycin and chloramphenicol were lower (11 and 9%, respectively). Five serotypes (6B, 1, 19A, 23F and 6A) accounted for 37% of the total isolates. Ten isolates (5%) were non-typeable. Overall, 29 and 42% of serotypes were represented in the 7- and 11-valent conjugate vaccines, respectively. However, vaccine coverage for children <2 years was 38 and 56% for the 7- and 11-valent, respectively. CONCLUSIONS: Resistance to penicillin may be increasing among S. pneumoniae strains causing meningitis in Egypt, and a moderate proportion of these strains are not covered by current pneumococcal conjugate vaccines. In addition to intensifying education efforts about judicious use of antibiotics, laboratory-based surveillance for other forms of invasive pneumococcal disease, especially pneumonia, is needed before decisions can be made regarding the most effective vaccines for control of this disease in Egypt.