Outbreak of foodborne botulism in Alexandria, Egypt: modulating indications for administration of heptavalent botulinum antitoxin

Sara A Ghitani Faculty of Medicine of Alexandria University: Alexandria University Faculty of Medicine Maha A Ghanem Faculty of Medicine of Alexandria University: Alexandria University Faculty of Medicine Eman A Sultan Alexandria Medicine: Alexandria University Faculty of Medicine Maram Atef (  maramatef99@gmail.com ) Alexandria University Faculty of Medicine https://orcid.org/0000-0002-4646-1326 Maii F Henaidy Faculty of Medicine of Alexandria University: Alexandria University Faculty of Medicine


Introduction
Botulism is considered one of the recurrent events that confront poison centers, in Egypt, each year due to a traditional salted sh meal which is consumed each spring (Weber et al.1993). In the spring of 2019, there were 58 cases of botulism admitted to APC and only 3 cases needed botulinum antitoxin. However, they suffered from side effects where 2 cases had severe epistaxis and one case arrested during antitoxin administration. On 29 September, a child was transferred to APC for differential diagnosis of paralytic ileus as her family said she was given a piece of salted sh (feseekh) and she was considered the rst case of the outbreak. Preliminary assessment of the situation was carried out and preparations were made to confront a large outbreak out of season.
A collaborative team from APC doctors with Egyptian Ministry of Health (EMOH) was initiated to conduct interviews with the cases about the source, the time of the start of symptoms, other personnel of same family exposed to same meal…. etc. Blood specimens were collected from the cases. Identi ed factors common to all cases and formulated preliminary hypotheses were issued. The team delivered a daily report to the authorities, and all types of media were used to issue a warning until the outbreak was stopped on 27-10-2019. The Egyptian news declared previously the death of almost 100 tons of "mullet" sh and veterinarians warning of the mass death of sh in a small town called Rashid which was considered the source of that outbreak (Veterinarians warn of the mass death of sh in Lake Rashid.2019).
Due to delay in results of mouse lethal test (MLT) and its positive and negative fallacies, the APC team treated the cases depending on the clinical picture (Lindstrom et  The current study aimed to portray the clinical characteristics of the cases, to explore the possible relation between these characteristics and necessity of HBAT administration, to explore the reliability of MLT and to establish a clinical guide of management of the suspected cases of botulism with preservation of resources.

III. Research tools:
A checklist and record review were carried out for collection of data to assess and follow up each patient.
The equine Heptavalent Botulism Antitoxin (A, B, C, D, E, F, G), made by Emergent BioSolutions Canada Inc. (formerly Can gene Corporation) was used in treatment. (8) Mouse Bioassay for Clostridium botulinum toxin was performed following FDA's Bacteriological Analytical Manual.

IV. Categorization of patients:
Clinical assessment was completed to all cases and a checklist including different signs and symptoms of botulinum toxicity was ful lled on admission.
To properly manage the resources, the patients were classi ed into the groups according to their clinical assessment that was repeated everyone hour. If the signs and symptoms were improving, assessment follow up would be every 6 hours.
The patients' groups were as the following: Group A: included those with history of Feseekh ingestion and were completely normal on their rst assessment and throughout their hospital stay (3 cases).
Group B: included patients complained of typical gastroenteritis throughout their course i.e., nausea, vomiting, diarrhea, abdominal pain, and mild abdominal distention (38 cases).
Group C: included all cases who developed neurological or autonomic dysfunction symptoms whether GIT symptoms were present or not. They were further subdivided into two groups: 1. Group C1: those at time of admission presented with evident serious neuromuscular dysfunction signs as ptosis, peripheral muscle weakness or hypoventilation (20 cases). The data were collected and stored into a computer. Statistical analysis was done using Statistical Package for Social Sciences (SPSS/version 20) software. Data was presented as numbers and percentages for categorical variables and mean ± standard deviation (SD) for continuous variables. For testing associations between qualitative variables, the chi square test was used. Signi cance of the obtained results was judged at the 5% level.

I. Demographic data:
The highest percentage of the participated cases (42.6%) were in the age group 20 -< 40 years. This was followed in a descending manner by the age group <20 years (34%), then the age group of 40-<60 years (21.3%) and the age group ≥ 60 years (2.1%). More than two thirds (69.1%) were females while 30.9% were males. As regard residence, 36.2% of the cases were from Alexandria, 35.1% came from Kafr El-Sheikh, 27.6% were from Elbehira and only one case (1.1%) was living in Al-Ismailia.

Past medical history:
In the current study, medical history of previous diseases, injuries and operative procedures were taken from all participants. However, most of the studied patients (91.5%) had no history of chronic diseases.
History of salted sh (Feseekh) ingestion: All the participated cases presented to the hospital from six hours to nine days after Feseekh ingestion with a mean value of 1.70 ± 1.49 days.
Assessment of the cases: The most common ndings observed in admitted patients were nausea and vomiting (76.6%), abdominal distension (41.5%) and dilated xed pupil (29.8%). (Table 1) Administration of HBAT: All cases with serious evident neurological signs either at time of admission or during observation received HBAT immediately. They were Thirty-four patients (36.2%). The rst dose of HBAT was given through 0.33 -6 days from Feseekh intake. Among those who received the rst dose of HBAT, more than half (55.9%) of the cases improved completely. On the other hand, six patients (17.7%) showed no improvement with persistent symptoms and 9 cases (26.4%) showed temporary improvement followed by relapse. Those fteen patients were given the second vial of the HBAT. The time interval between the rst and second vials was 1-4 days with a mean of 2.03± 0.93 days. Only one patient needed a third vial of the botulinum antitoxin. This was attributed to the persistence of hypoventilation one day after the second dose.
The patients in group C2 were further categorized according to the number of mild cranial de cit complains and the need for HBAT administration and the results revealed that: All patients who admitted to hospital complaining of only one symptom of autonomic dysfunction did not need the antitoxin, while 100% of patients presented with four autonomic symptoms needed the antitoxin during their period of observation. On the other hand, 77.8% of those with three autonomic dysfunction symptoms needed the antitoxin. However, half of those with only two autonomic symptoms developed neurological signs later and needed the antitoxin. And this difference is statistically signi cant with p< 0.001. (Table 2) 6. Result of mouse bioassay test: A blood sample was withdrawn from patients on admission to the poison center even after days of ingestion time of the Feseekh and serum was prepared and transferred to the lab. Mouse bioassay test was carried out to 82 out of 94 cases and 87.8 % was negative. It means that 24 cases with negative mouse bioassay test developed neurological symptoms and needed HBAT. (Table 3) 7. Duration of hospitalization, ICU admission and need to mechanical ventilation support: The duration of hospital admission was 1-12 days with a mean of 3.71 ± 2.75 days. Moreover, 30 patients (31.9%) admitted to ICU for a period ranged from 1-8 days with a mean of 3.40 ± 1.81 days.
Eight patients (26.7%) from group C1 needed mechanical ventilation for a period of 1-5 days with a mean of 2.50 ±1.60 days.
8. The outcome: Recovery was the major outcome (87.2%) whereas neurological sequalae were present in 10.6% of patients -in the form of persistent constipation, distention, dry mouth with a very mild dysphagia and death occurred only in two cases (2.2%).

Discussion
On spring, Egyptians' celebration of eastern festival is accompanied with eating Feseekh (salted sh).
However, unexpected outbreak occurred in Autumn and it was justi ed by contamination of marine mullet meat during that time, where some of those contaminated sh were taken by some individuals and salted in their homes. (Veterinarians warn of the mass death of sh in Lake Rashid 2019).
The outbreak started with a 1.5-year-old child referred from Pediatric University Hospital to APC for differential diagnosis. She presented with paralytic ileus that raised the suspicion of intussusception and surgical abdomen. On examination the surgical emergency was eliminated. A similar scenario was mentioned by Spini et al (2015).
Due to the unusual young age of the child with the absence of the common paralytic manifestations of botulism, pediatricians doubted botulism. Pisani et al (2009) had explained a similar situation as the initial symptoms of the disease are often like several diseases and therefore differential diagnosis is very di cult and rarely suspected by the pediatrician.
Although the highest percentage of the patients were in the middle age group (20 -< 40 years), extremes of age were also involved, and females outnumbered males. This could be explained by the Egyptian culture where the ladies prepared the meals to the families as well as tasting it before meals. All family members gathered for celebration and ate the same food so different ages are usually affected.
APC is serving Alexandria and all-around governorates (Elbehira, Kafr el-Sheikh, Marsa Matrouh, etc.). As the accident of contamination of the mullet sh meat occurred in Elbehira governorate so, all patients were directed to APC as in all other poisons.
In the current work, 91.5% of the cases had no previous medical or surgical history. Past medical and surgical history was taken to exclude neurological diseases, previous stroke, and other gastrointestinal problems. All patient's symptoms started to appear after eating the Feseekh meal. All the participated cases presented to the hospital from six hours up to nine days after Feseekh ingestion with a mean of Although the clinical syndrome of botulism is distinctive, it needs a high index of suspicion to be diagnosed. In the current study, patients were presented with mydriasis (29.8%), blurred vision (14.9%), ptosis (11.7%), dysphagia (24.5%) and muscle weakness in 21.3% of patients. This is in accordance with Gaware et al (2011) who reported these symptoms as the classic for botulism.
On contrary to most of previous studies on Botulism that recommend early antitoxin administration to reduce mortality Gaware et al (2011), Kongsaengdao et al (2006) and Yu (2015). The current study aimed to manage the resources and to decrease the exposure of the patients to the side effects of the unnecessary administration of antitoxin. Thus, the patients in current study were classi ed at the time of admission into three groups. Only twenty patients -group C1-who presented with severe evident neurological symptoms; ptosis, muscle weakness or respiratory muscle affection with hypoventilation were given the BAT immediately. While the rest of patients didn't receive the BAT immediately but they were put under close observation.
Patients in group A who presented to hospital with just a history of Feseekh ingestion as members of their family ate the same meal and developed botulism manifestations and also patients in group B who presented with only gastroenteritis didn't develop any additional symptoms during their period of hospitalization and did not need to receive the BAT. Moreover, patients in group C2 who presented with mild cranial de cit didn't receive the BAT immediately as most of cranial de cits complain of patients in group C2 might be just a subjective feeling due to the fear or anxiety that result from the warnings spread at that time on the media or due to their affection by other patients in the outbreak.
However, in order to identify patients likeliest to bene t from the antitoxin treatment, patients in group C2 were categorized according to the number of cranial de cit signs on initial assessment to anticipate the deterioration and to predict who would need the antitoxin administration which may be valuable for further development of guidelines. The initial assessment of cases with suspected food-born botulism is very important.
In our opinion, this classi cation reduced the unnecessary administration of the HBAT to 60 patients who were managed by supportive treatment only. This protocol helped avoidance of many side effects of the antitoxin including the most severe ones as anaphylaxis and hypersensitivity reactions and other delayed allergic reactions as serum sickness. Schussler et al (2017)  The only drawback of the current protocol is prolongation of hospitalization due to monitoring of the patients which reaches up to 12 days in some cases to avoid missing the unpredicted side effects of the antitoxin.
Following the theory in current study, complete recovery occurred in (87.2%). Complications were present in 10.6% of patients while death occurred only in two cases (2.2%). All those 10.6% of patients who discharged with complications received the HBAT. They complained of persistent dry mouth with a very mild dysphagia not interfering with uid, solid or semisolid food ingestion. Some complained of constipation which improved also with the usage of laxatives. This is like Gottlieb et al (2007) who concluded that previous symptoms of cholinergic autonomic blockade that are prominent during botulism as dry mouth would persist for a long-term later.
Regarding the two cases who died, both were in group C1 with evident neurological paralysis, they received the antitoxin, and both were in need for mechanical ventilation in ICU. They arrested in the ICU; one after 2 days and the other after 4 days without receiving any further doses of HBAT. According to the critical care physicians the cause of death was a superadded chest infection. This point is very important as the appropriate supportive care is still the cornerstone in recovery and survival after botulism. As those cases suffered respiratory compromise so a high-quality intensive care is needed. This is in accordance with Sheth et al (2008).

Conclusions
Evidence based from that research that the course of cases with suspected botulism often has a stationary course and 63.8% of cases could be managed by supportive treatment with no need for HBAT, so we advise to keep the patient under close observation instead of rushing to give the anti-botulinum antitoxin, so we can protect him from the side effects of HBAT and manage resources at the same time especially in outbreaks. However, in evident neurological manifestations HBAT administration should be administered immediately.

Declarations
Author's contribution: Sara A ghitani participated in the design of the study, drafted the manuscript, and participated in reviewing its nal version, Maha A Ghanem participated in data collection and in drafting the manuscript and reviewing the nal version, Eman A Sultan participated in the design of the study and performed the statistical analysis, Maram Atef participated in data collection and storage and reviewing the nal version and Maii F Henaidy participated in writing the results section.