|Year : 2021 | Volume
| Issue : 1 | Page : 39-45
Prevalence and risks factors of the novel Escherichia albertii among gastroenteritis patients in Kano State, Nigeria
Mamunu A Sulaiman, Maryam Aminu, Elijah E Ella, Isa O Abdullahi
Department of Microbiology, Faculty of Life Sciences, Ahmadu Bello University, Zaria, Nigeria
|Date of Submission||03-Jun-2020|
|Date of Decision||02-Jul-2020|
|Date of Acceptance||30-Sep-2020|
|Date of Web Publication||28-Apr-2021|
Mamunu A Sulaiman
Department of Microbiology, Faculty of Life Sciences, Ahmadu Bello University, Zaria
Source of Support: None, Conflict of Interest: None
Background: Escherichia albertii is an enteric bacteria associated with infectious gastroenteritis. However, information on this organism remains scant in Nigeria. The work was aimed at determining the prevalence of E. albertii among gastroenteritis patients (GEP) in relation to some risk and demographic factors. Methods: The study was a cross-sectional hospital-based research. A total of 540 stool samples, 450 from GEPs and 90 from apparently healthy individuals, were screened for the pathogen. The isolates were characterized by using both conventional and molecular techniques. Data were obtained using a structured questionnaire and were analyzed for the determination of some risk and demographic factors. Odds ratio and chi-square were used to ascertain the relationship between the factors and the disease. Results: Escherichia albertii was detected with an overall prevalence of 1.1%, which was 1.3% and 0% in the GEPs and apparently healthy individuals, respectively (Odd ratio = 2.6468, 95% CI, 0.1478–47.4042). Diarrhea and abdominal pain were statistically the significant manifestations of the disease. There was an equal prevalence in both males and females (1.3%), however, by chance higher (2.1%) among children aged 0 to 10 years. Additionally, primary school as the highest educational status and open defecation were found to be the risk factors of the infection. Conclusions: Escherichia albertii should be routinely targeted diagnostically in the patients presented with gastroenteritis in the studied area. Neither gender nor age was a risk factor, however open defecation was a significant risk factor of the disease.
Keywords: Demographic factors, gastroenteritis, Kano State, Nigeria, risk factors
|How to cite this article:|
Sulaiman MA, Aminu M, Ella EE, Abdullahi IO. Prevalence and risks factors of the novel Escherichia albertii among gastroenteritis patients in Kano State, Nigeria. J Med Trop 2021;23:39-45
|How to cite this URL:|
Sulaiman MA, Aminu M, Ella EE, Abdullahi IO. Prevalence and risks factors of the novel Escherichia albertii among gastroenteritis patients in Kano State, Nigeria. J Med Trop [serial online] 2021 [cited 2021 Sep 19];23:39-45. Available from: https://www.jmedtropics.org/text.asp?2021/23/1/39/314851
Key Messages: Escherichia albertii was reported with a prevalence of 1.3% among the gastroenteritis patients in Kano State, Nigeria. The infection may be more common in a population where open defecation is practiced, and may likely be manifested as diarrhea with abdominal pain.
| Introduction|| |
Infectious gastroenteritis is an important cause of morbidity and mortality, especially in the environment with poor sanitary conditions., The genus Escherichia is one of the most important enteric bacteria, composed of five species, Escherichia coli and four less frequently encountered members that are Escherichia hermannii, E. vulneris, E. fergusonii, and E. blattae. Subsequently, the sixth species, Escherichia albertii was isolated associated with diarrheal disease in Bangladesh children . Escherichia albertii are transmitted through contaminated water or food, making them a threat to public health, and have been added to the group of attaching and effacing pathogens. Escherichia albertii are Gram-negative, non-spore formers, and non-lactose fermenters but ferment d-glucose (with both acid gas production). They are non-motile and lack the inability to ferment xylose or dulcitol., Their ability to ferment sucrose is variable. They are methyl red positive, and Voges–Proskauer (VP) negative. They produce acid from the fermentation of d-mannitol, d-mannose, and galactose, and reduce nitrate. A high percentage (73.7%–97.4%) of the organism is positive for the utilization of ortho-nitrophenyl-B-galactoside (ONPG), trehalose, lysine, glycerol, and acetate. Variable positive reactions (35%–44.7%) were documented for the reaction of the organism to indole and d-sorbitol tests. The majority of the isolates do not ferment raffinose, l-rhamnose, or utilize citrate., To the best of our knowledge, currently there were no published data on Escherichia albertii from Nigeria or even the entire West Africa. The worldwide report on this pathogen is still scant, confounding the scientific understanding of E. albertii, their role in causing diseases in humans and animals, and in using rapid kits to characterize them. The aim of the work was to determine the prevalence of E. albertii among gastroenteritis patients (GEPs) in relation to some risk and demographic factors.
| Materials and methods|| |
The samples were collected from six hospitals across the three senatorial zones of Kano State and transported in peptone water. Kano State is having a population of about 9,383,682 in the 2006 census. It has 44 local governments, with an area of 20,479.6 square kilometers. Kano is 481 meters above sea level.
This research was a hospital-based cross-sectional study. Ethical approval was obtained from Kano State Hospitals Management Board prior to the commencement of the study. Similarly, an informed consent was obtained from the patient/guardian/caregiver in case of children prior to sample collections from the subjects. A structured questionnaire was used, to capture data on possible risk and socio-demographic factors.
Children and adults (irrespective of the gender) with suspected cases of gastroenteritis, presented with diarrhea in combination with fever, abdominal pain, nausea, or abdominal cramps attending Gwarzo General Hospital, Bichi General Hospital, Muratala Mohd General Hospital, Abdullahi Wase Special Hospital, Gaya General Hospital, and Sumaila General Hospital were included.
Individuals with no apparent symptoms of gastroenteritis and those that did not consent.
The sample size was calculated according to the formula of Zubair; the prevalence used was 38.9%.
A total of 540 stool samples (450 from GEPs and 90 from AHIs) were collected from six hospitals, distributed across the three senatorial zones of Kano State.
The stool samples were physically examined for the formation presence of blood or mucus. The formation of the stool was categorized as watery if accompanied with a copious quantity of water, semi-formed if appeared semi-solid, or formed if the texture was normal. Additionally, the presence of blood or mucus was used to categorize stool as bloody or mucoid if visibly observed.
Cultural isolation and characterization of E. albertii
The samples were streaked aseptically on MacConkey agar plates and incubated overnight at 37°C. The colonies that appeared as non-lactose fermenting, non-mucoid were considered for the biochemical characterization.
Conventional characterization of E. albertii
Gram staining technique was used for the preliminary identification. Motility of the organism was tested along with H2S and indole production using sulfide indole motility (Oxoid) medium slant. Additionally, methyl red (MR) and Voges–Proskauer (VP) tests were carried out using MR-VP broth (Oxoid). Also, d-glucose, lactose, and sucrose fermentation tests were carried out using triple sugar iron (TSI) medium (Oxoid). Also, sugar fermentation test was carried out by preparing 1% of each of the d-arabitol, dulcitol, l-rhamnose, raffinose, d-sorbitol, d-mannitol, d-xylose, and l-arabinose in peptone water. Lastly, decarboxylation of the isolates was tested against lysine and ornithine. All the biochemical tests were carried out under overnight incubation at 37°C in accordance with the manufacturer’s instruction.
Molecular characterization of E. albertii
The genetic material was extracted from fresh cultures, grown overnight in nutrient broth, and then centrifuged (Bio-Rad centrifuge) at 10,000 rpm for 10 minutes. The supernatants were decanted and the sediments were suspended in 0.5 mL of 1× PBS and centrifuged again for another 10 min at 10,000 rpm. The supernatants were decanted and the sediments were re-suspended in 0.5 mL of sterilized distilled water. The cells were lysed by incubation in the heating block at 100°C for 10 minutes after votexing. The tubes were kept in ice for 15 minutes and then centrifuged at 16,000 rpm for 10 minutes. Then 0.5 mL of the supernatants (DNA) was pipetted and stored at −2°C until further analysis.
Multiplex PCR for confirmation of E. albertii
Each reaction was prepared as 25 μL, which contained 12.5 μL of the master mix (Biolabs, England), 3 μL of the DNA, 0.5 μL (10 μmol) each of the forward and reverse primers (Inqaba BioTech), and 7.5 μL molecular-grade water. The PCR was run using a duo of EA primer and EC primers synthesized at Inqaba Biotec Inc (South Africa; [Table 1]). The multiplex PCR was run using a thermocycler (BioRad, USA): initial denaturation at 94°C for 30 seconds followed by subsequent denaturation at 94°C for 30 seconds, annealing at 48°C for 1 minute, and extension at 68°C for 1 minute. The condition (except the initial denaturation) was run for 35 cycles before the final extension at 68°C for 5 minutes.
|Table 1: Primer sequences of genes for the characterization of E. albertii using multiplex PCR.|
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The PCR products were cast in a 1.5% agarose gel with ethidium bromide (5 μL/100 mL) as the intercalating dye and then set at 80 V for 40 minutes. The amplicons were viewed and documented using UV light imaging system (BioDoc-It™).
Odds ratio/chi-square was calculated using SPSS software (IBM, 22.0) to ascertain the level of statistical association or differences within the categories.
| Results|| |
E. albertii are gram-negative bacilli, non-motile, and H2S and indole negative. They were negative to both MR and VP and ferment only d-glucose in TSI medium (K/A). Additionally, the pathogen was characterized as non-fermenter of all the sugars tested in the study except d-sorbitol and d-mannitol. However, the isolates decarboxylated both lysine and ornithine. The isolates characterized biochemically as E. albertii were confirmed using the molecular technique.
Six isolates from GEPs were confirmed as E. albertii, showing both ec (212bp) and ea (393bp) genes as observed on lanes 7 and 17 [Plate 1] and lanes 2, 3, 7, and 17 [Plate 2], with research codes: AG109, BG253, YG309, SG381, SG391, and SG431, respectively. Thus, the overall prevalence of E. albertii is 1.1%; 1.3% and 0% among the GEPs and apparently healthy individuals (AHIs), respectively (odds ratio = 2.6468, 95%CI, 0.1478–47.4042; [Table 2]). The highest (3.0%) and lowest (0%) prevalence of E. albertii was recorded in patients with diarrhea and formed stool, respectively, with the differences observed statistically significant (P = 0.01975). Diarrhea was mostly (7.7%) accompanied by blood, compared with the non-bloody diarrhea (0.95%), and the differences observed were by chance (P = 0.0723). Also, the prevalence was higher (1.9%) among those who experienced constipation and lower (1.3%) among those who did not, the difference was by chance (P = 0.6934). Also, the highest (7.9%) and lowest (0.5%) prevalence was observed among the patients with abdominal pain and those who did not experience any of the localized symptoms, respectively, and the differences were statistically significant (P = 0.0026). Similarly, the highest prevalence was observed among the patients who experienced fever (4.8%) compared to the lowest (0.4%) among those who did not, and the differences observed were statistically insignificant (P = 0.0716; [Table 3]). The prevalence of E. albertii in males (2/153) and females (4/297) was the same (1.3%). Also, the prevalence was higher (2.1%) in the age group of 0 to 10 years old primary school pupils, followed by 1.2% in the age group of 11 to 20 years old, and 0% down the rest of the age groups, and the differences observed were statistically insignificant (P = 0.7276). With regards to the level of education, the highest prevalence was observed among primary school children and the lowest of 0% each among secondary school and tertiary institution students, and the differences observed were statistically significant (P = 0.0109). The highest (1.9%) prevalence was among those who were using well water as the major source for drinking and lowest among those who were using sachet water (1.2%), and the differences observed were statistically insignificant (P = 0.8821; [Table 4]).
|Table 2: Prevalence of Escherichia albertii among gastroenteritis patients and apparently healthy individuals in Kano State, Nigeria|
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|Table 3: Prevalence of E. albertii in relation to some symptoms and medical history of the gastroenteritis patients|
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|Table 4: Prevalence of E. albertii in relation to some demographic and risk factors of the gastroenteritis patients|
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| Discussion|| |
From our findings, E. albertii was approximately 2.6 times more common among the GEP than AHI. The current report is in agreement with the previous study that reported E. albertii as an agent of gastroenteritis outbreak in Japan with 100% prevalence. The much higher prevalence in the previous study maybe because all the subjects got exposed accidentally to the same source of infection (contaminated food).
In the current study, diarrhea is a significant symptom of E. albertii-associated gastroenteritis and it may be accompanied by blood or alternates with constipation. This pathogenesis may be due to the virulence factors as stxI and II and eae harbored by the pathogen, which are known to interact with the human intestine with resultant fluid accumulation that could result to diarrhea and abdominal pain as observed in the study. Also, the products of these genes have been implicated in the destruction of the intestinal villi and that may be the cause of the blood observed in diarrhea. Additionally, the virulence factors have been implicated in electrolyte imbalance, which may be the cause of constipation. The toxins may ulcerate the intestine, giving the pathogen a route of propagation and spreading to the circulatory system, with the outcome observed as systemic symptoms inform of fever, headache, or both. From the statistical observation in the current study, fever could have happened in the patients by chance, but clinically could be an indication of having a bacteremic stage during the infection, however, abdominal pain was statistically proven diagnostic of the infection. Both the abdominal pain and the fever were previously reported among people who got infected with E. albertii after attending a party in a restaurant (in Japan) with the prevalence of 76% and 38%, respectively .
The prevalence of E. albertii was equal among both males and females, ruling out gender as a possible risk factor of its gastroenteritis. The result is in agreement with the previous work, where gender was similarly reported as an insignificant risk factor of infectious gastroenteritis. The prevalence of E. albertii gastroenteritis reduces drastically by almost half from the age groups 0 to 10 years to 11 to 20 years, and then completely to 0% in the age group (years) 21 to 30, 31 to 40, and >40. Although there was no significant statistical association in the observed differences, but this is an indication that younger children are at higher risk of the infection than adults, which was in agreement with the first scientific report of the pathogen. The predilection of E. albertii for children in our findings explains the significance of their infection among primary school children, and the school as the transmission ground. None of the sources of drinking water was statistically implicated for E. albertii associated gastroenteritis, however, well water conferred a higher risk of getting the infection, and borehole water was the safest. Moreover, the organism was reported as a resilient contaminant of pipe-borne water. The level of awareness of the mode of transmission of infectious diseases is still less than enough, especially villages of many underdeveloped countries like Nigeria. This suggests why people in such areas are lagging in taking precautionary hygienic and sanitary measures necessary for the prevention and control of such diseases. From our findings, lack of awareness of the mode of the transmission of the pathogen was not a risk factor, as such, it is in contradiction to some reports on some enteric pathogens. Open defecation was a significant risk factor in acquiring E. albertii-associated gastroenteritis, and water closet is the safest type of toilet. Good hand wash practices can prevent infectious diseases transmitted through fecal–oral route by up to 95%, however, the level of the prevention reduces if it is poorly practiced, perhaps why it was not preventive in the current study, as there was no statistical evidence to implicate non-practicing hand wash after toilet and before eating as a risk factor of E. albertii infection. The result is in contradiction with the previous work, where poor hand wash practices ware reported as statistically significant factors, associated with the rate of fecal borne infection, in some parts of Kaduna state.,
| Conclusion|| |
The prevalence of E. albertii from stool samples of GEPs and AHIs was 1.3% and 0% respectively. The infection was presented with diarrhea and abdominal pain, and these can be considered in clinical diagnosis. The infection was most common among primary school children and open defecation was implicated as the risk factor of the infection.
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Conflicts of interest
There are no conflicts of interest.
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[Plate 1], [Plate 2]
[Table 1], [Table 2], [Table 3], [Table 4]